Aah1 is a GPI-anchored, cell-surface GH13-family enzyme that functions as an alpha-glucan transglycosylase (4-alpha-glucanotransferase, EC 2.4.1.25) in S. pombe. It acts redundantly with its paralog Aah3 to remodel alpha-glucan chains synthesized by the alpha-glucan synthase Ags1, building a properly organized alpha-1,3-glucan matrix in the cell wall. Loss of both aah1 and aah3 causes severe reduction of alpha-1,3-glucan, rounded cell morphology, delayed cell division, clumping, and cell wall thickening with compensatory beta-glucan increases. Single aah1 deletion has no obvious phenotype under standard conditions, indicating functional overlap with aah3.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
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GO:0004134
4-alpha-glucanotransferase activity
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IEA
GO_REF:0000003 |
ACCEPT |
Summary: This IEA annotation from EC mapping to EC 2.4.1.25 is consistent with the UniProt-assigned EC number and with the strong functional inference from close A. niger orthologs (AgtA/AgtB) that are biochemically characterized as 4-alpha-glucanotransferases.
Reason: The EC 2.4.1.25 assignment is well supported by domain conservation, catalytic triad preservation, and direct enzymology of GPI-anchored GH13 fungal orthologs. The IEA annotation is redundant with the ISS/ISO annotations below but consistent.
Supporting Evidence:
PMID:40668835
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly with the α-glucan synthase to build a properly organized α-glucan matrix.
file:SCHPO/aah1/aah1-deep-research-falcon.md
Two GPI-anchored A. niger GH13 enzymes (AgtA/AgtB) were biochemically shown to have 4-alpha-glucanotransferase activity (EC 2.4.1.25), cleaving alpha-(1,4) bonds and forming new alpha-(1,4) linkages.
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GO:0005509
calcium ion binding
|
IEA
GO_REF:0000002 |
KEEP AS NON CORE |
Summary: Calcium ion binding is inferred from the InterPro alpha-amylase C-terminal domain (IPR015340). UniProt notes by similarity that Aah1 binds 2 calcium ions per subunit, based on conservation with alpha-amylase family members. This is a structural/cofactor role common to GH13 enzymes.
Reason: Calcium binding is a conserved structural feature of GH13 enzymes required for proper folding and catalytic activity, but it is not the core function of Aah1. The annotation is likely correct based on domain conservation but represents a cofactor requirement rather than the primary molecular function.
Supporting Evidence:
file:SCHPO/aah1/aah1-uniprot.txt
Binds 2 calcium ions per subunit. Calcium is inhibitory at high concentrations.
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GO:0005886
plasma membrane
|
IEA
GO_REF:0000044 |
KEEP AS NON CORE |
Summary: Aah1 is a GPI-anchored protein tethered to the plasma membrane on the extracellular side. While technically attached to the plasma membrane, the more informative localization is the external side of the plasma membrane / fungal-type cell wall where it carries out its function.
Reason: The plasma membrane annotation is correct but less informative than the external side of plasma membrane (GO:0009897) annotation that is also present. The GPI anchor tethers the protein to the membrane, but the functional localization is extracellular/cell wall.
Supporting Evidence:
PMID:40668835
...two conserved glycosylphosphatidylinositol-anchored α-amylase-like enzymes, Aah1 and Aah3, which act redundantly as key contributors to α-glucan network formation.
file:SCHPO/aah1/aah1-uniprot.txt
Cell membrane; Lipid-anchor, GPI-anchor; Extracellular side.
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GO:0005975
carbohydrate metabolic process
|
IEA
GO_REF:0000002 |
MARK AS OVER ANNOTATED |
Summary: This is a very broad parent term for the more specific cell wall polysaccharide biosynthetic processes that Aah1 is involved in. The annotation is technically correct but overly general.
Reason: The specific biological processes GO:0070600 (fungal-type cell wall (1->3)-alpha-glucan biosynthetic process) and GO:0051278 (fungal-type cell wall polysaccharide biosynthetic process) are more informative and already present. This broad parent term does not add useful information.
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GO:0009897
external side of plasma membrane
|
IEA
GO_REF:0000117 |
ACCEPT |
Summary: Aah1 is a GPI-anchored protein localized to the external side of the plasma membrane where it acts on cell wall alpha-glucans. This is consistent with the GPI anchor prediction and the functional role of remodeling extracellular alpha-glucan polymers.
Reason: The external side of the plasma membrane is the correct and informative localization for a GPI-anchored cell surface enzyme. Supported by the GPI-anchor signal in the sequence and the known cell-wall-associated function.
Supporting Evidence:
PMID:40668835
...two conserved glycosylphosphatidylinositol-anchored α-amylase-like enzymes, Aah1 and Aah3, which act redundantly as key contributors to α-glucan network formation.
PMID:16751704
...Aah3p is an N-glycosylated, GPI-anchored membrane protein localized in the membrane and cell wall fractions.
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GO:0016052
carbohydrate catabolic process
|
IEA
GO_REF:0000002 |
REMOVE |
Summary: This annotation from InterPro IPR015340 (alpha-amylase C-terminal domain) suggests a catabolic role. However, Aah1 functions as a transglycosylase rather than a hydrolase. No starch-degrading activity was detected in S. pombe cell lysates or culture media. The protein remodels cell wall alpha-glucans rather than degrading carbohydrates.
Reason: Aah1 is not involved in carbohydrate catabolism. It is a transglycosylase that remodels alpha-glucan polymers in the cell wall. No hydrolytic/catabolic activity was detected experimentally, and the protein's biological role is biosynthetic/remodeling rather than degradative.
Supporting Evidence:
PMID:16751704
Expression of aah3 alleles in which the conserved aspartic acid and glutamic acid residues required for hydrolase activity were replaced with alanine residues failed to rescue the morphological and ascal wall defects of aah3Delta cells.
PMID:40668835
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly with the α-glucan synthase to build a properly organized α-glucan matrix.
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GO:0004134
4-alpha-glucanotransferase activity
|
ISS
GO_REF:0000024 |
ACCEPT |
Summary: ISS annotation based on manual transfer from ortholog A2QTS4 (A. niger AgtA). AgtA was biochemically demonstrated to have 4-alpha-glucanotransferase activity on alpha-(1,4)-linked glucan donors requiring at least ~5 glucose units, with maltose as the smallest effective acceptor.
Reason: This is the core molecular function of Aah1. The ISS transfer from the biochemically characterized A. niger ortholog is well justified given the conserved GH13 catalytic domain, GPI-anchor architecture, and the genetic evidence from PMID:40668835 supporting a transglycosylase role in cell wall alpha-glucan remodeling.
Supporting Evidence:
PMID:40668835
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly with the α-glucan synthase to build a properly organized α-glucan matrix.
file:SCHPO/aah1/aah1-deep-research-falcon.md
Two GPI-anchored A. niger GH13 enzymes (AgtA/AgtB) were biochemically shown to have 4-alpha-glucanotransferase activity. Their reactions are dominated by transglycosylation: they cleave alpha-(1,4) bonds in donor substrates and form new alpha-(1,4) linkages in products.
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GO:0004134
4-alpha-glucanotransferase activity
|
ISO
GO_REF:0000024 |
ACCEPT |
Summary: ISO annotation from PomBase, also based on orthology to A2QTS4. This is the same functional assignment as the ISS annotation above but from a different source (PomBase vs UniProt).
Reason: Consistent with the ISS annotation and well supported by the same evidence. The ISO evidence code reflects orthology-based transfer, which is appropriate given the high domain conservation and functional similarity demonstrated by genetics.
Supporting Evidence:
PMID:40668835
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly with the α-glucan synthase to build a properly organized α-glucan matrix.
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GO:0051278
fungal-type cell wall polysaccharide biosynthetic process
|
ISS
GO_REF:0000024 |
ACCEPT |
Summary: ISS annotation transferred from ortholog. Aah1 contributes to biosynthesis of cell wall alpha-glucan polysaccharides. The aah1/aah3 double mutant shows dramatic reduction in cell wall alpha-1,3-glucan (from 44% to 11% in rigid phase).
Reason: Aah1 is directly involved in building the alpha-glucan component of the fungal cell wall. While the single mutant has no clear phenotype, the double mutant with aah3 demonstrates that Aah1 redundantly contributes to cell wall polysaccharide biosynthesis.
Supporting Evidence:
PMID:40668835
...the double mutant cell walls have dramatically reduced α-1,3-glucan and galactomannan content, with a compensatory increase in β-glucans driven by the activation of the cell integrity pathway.
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GO:0071940
fungal-type cell wall assembly
|
ISS
GO_REF:0000024 |
ACCEPT |
Summary: ISS annotation for cell wall assembly. Aah1 contributes to proper cell wall architecture by remodeling alpha-glucan chains into mature organized fibers. The double mutant has severely altered wall structure with 4-fold thickening.
Reason: Cell wall assembly is a core biological process for Aah1. The protein remodels alpha-glucan polymers synthesized by Ags1 into a properly organized matrix, which is essential for normal cell wall architecture, rod shape, and cell separation.
Supporting Evidence:
PMID:40668835
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly with the α-glucan synthase to build a properly organized α-glucan matrix... Cells lacking both enzymes exhibit severe growth and morphological defects, including rounded shape, delayed division, and cell clumping.
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GO:0070600
fungal-type cell wall (1->3)-alpha-glucan biosynthetic process
|
IMP
PMID:40668835 α-glucan remodeling by GH13-domain enzymes shapes fungal cel... |
ACCEPT |
Summary: IMP annotation from PomBase based on the Jacob et al. 2025 study. The aah1/aah3 double mutant shows dramatic reduction in alpha-1,3-glucan (from 44% to 11% in rigid phase). This is the most specific and informative biological process annotation.
Reason: This is the most specific and experimentally supported biological process annotation for Aah1. The IMP evidence from solid-state NMR analysis directly demonstrates that loss of Aah1 (with Aah3) leads to severe reduction in alpha-1,3-glucan in the cell wall. This represents the core biosynthetic process in which Aah1 participates.
Supporting Evidence:
PMID:40668835
...the double mutant cell walls have dramatically reduced α-1,3-glucan and galactomannan content, with a compensatory increase in β-glucans driven by the activation of the cell integrity pathway.
file:SCHPO/aah1/aah1-deep-research-falcon.md
Rigid-phase carbohydrate composition: alpha-1,3-glucan decreases from 44% to 11% in aah1/aah3 double mutant.
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GO:0009277
fungal-type cell wall
|
ISS
GO_REF:0000024 |
ACCEPT |
Summary: ISS annotation from PomBase for localization to the fungal-type cell wall. Aah1 is a GPI-anchored enzyme that localizes to the cell surface/cell wall where it remodels alpha-glucan polymers. This localization is consistent with the GPI-anchor signal and the known cell wall function.
Reason: As a GPI-anchored cell surface enzyme that remodels cell wall alpha-glucans, localization to the fungal-type cell wall is appropriate. The paralog Aah3 was shown to localize to membrane and cell wall fractions biochemically.
Supporting Evidence:
PMID:16751704
...Aah3p is an N-glycosylated, GPI-anchored membrane protein localized in the membrane and cell wall fractions.
PMID:40668835
...two conserved glycosylphosphatidylinositol-anchored α-amylase-like enzymes, Aah1 and Aah3, which act redundantly as key contributors to α-glucan network formation.
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Q: What is the direct enzymatic activity and substrate specificity of purified Aah1? No in vitro assay on purified Aah1 alone has been reported.
Q: Does Aah1 have any unique substrates or products compared to Aah3, given that they are described as acting nonredundantly with each other?
Q: What is the individual contribution of aah1 to cell wall alpha-glucan under stress conditions or during specific cell cycle stages?
Experiment: Purify recombinant Aah1 and characterize its substrate specificity and products using defined alpha-glucan oligosaccharides as donors and acceptors.
Experiment: Perform solid-state NMR analysis on aah1 single mutant (comparing to wild type and aah3 single mutant) to determine the individual contribution of Aah1 to cell wall composition.
The research report should be a detailed narrative explaining the function, biological processes, and localization of the gene product. Citations should be given for all claims.
You should prioritize authoritative reviews and primary scientific literature when conducting research. You can supplement
this with annotations you find in gene/protein databases, but these can be outdated or inaccurate.
We are specifically interested in the primary function of the gene - for enzymes, what reaction is catalyzed, and what is the substrate specificity? For transporters, what is the substrate? For structural proteins or adapters, what is the broader structural role? For signaling molecules, what is the role in the pathway.
We are interested in where in or outside the cell the gene product carries out its function.
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Include evidence where possible. We are interested in both experimental evidence as well as inference from structure, evolution, or bioinformatic analysis. Precise studies should be prioritized over high-throughput, where available.
The fission yeast S. pombe gene aah1 encodes a predicted cell-surface/cell-wall α-amylase–superfamily (GH13) protein with a C-terminal signal consistent with GPI anchoring and conserved catalytic residues typical of α-amylase–fold enzymes. Although direct in vitro enzymology for Aah1 alone was not recovered in the accessible literature here, strong genetic/biophysical evidence indicates that Aah1 functions in cell-wall α-glucan architecture. In particular, loss of Aah1 together with its paralog Aah3 causes major reductions in cell-wall α-1,3-glucan and α-1,4-glucan, severe cell-shape and separation defects, and compensatory increases in β-glucans. These phenotypes support a model in which Aah1 acts as a cell-wall α-glucan transglycosylase/remodeling enzyme, likely performing 4-α-glucanotransferase-like reactions (EC 2.4.1.25) on α-1,4-linked donors and producing or rearranging α-glucan segments to enable proper α-1,3-glucan fiber formation. (jacob2025solidstatenmranalysis pages 9-12, jacob2025solidstatenmranalysis pages 5-9, jacob2025solidstatenmranalysis pages 1-5)
Enzymes in the α-amylase superfamily (including GH13 and related GH77 transferases) act on α-glucosidic bonds and can partition between hydrolysis and transglycosylation via a retaining, double-displacement mechanism. A common catalytic framework includes an Asp nucleophile, a Glu acid/base, and a second Asp that stabilizes the transition state, embedded in the (β/α)8-barrel fold. (vosUnknownyearlossofglucosidasea pages 45-48)
A 4-α-glucanotransferase (EC 2.4.1.25; also called amylomaltase-like activity) is a transglycosylase that cleaves α-(1,4) linkages in a donor α-glucan and transfers the released glucosyl segment to an acceptor—often maltose or maltooligosaccharides—forming a new α-(1,4) linkage. This chemistry is established experimentally for fungal, cell-wall–associated, GPI-anchored GH13 enzymes (see §3). (kaaij2007twonovelputatively pages 1-2)
In S. pombe, the cell wall contains β-glucans and an essential α-glucan component; α-glucan synthesis is associated with the multidomain α-glucan synthase Ags1p, which includes an extracellular transglycosylase-like domain and an intracellular synthase domain. α-glucan remodeling enzymes also contribute to processes such as cell separation. (vosUnknownyearlossofglucosidase pages 183-189, vosUnknownyearevidenceforthea pages 183-189)
Peer-reviewed S. pombe work on the aah gene family (aah1–aah4) identifies Aah1 as an α-amylase-like protein that retains the canonical catalytic residues (Asp/Glu/Asp) and contains a C-terminal hydrophobic region consistent with a GPI-anchor signal. (Morita et al., 2006; publication date: 2006-06; URL: https://doi.org/10.1271/bbb.50693) (morita2006anαamylasehomologue pages 3-5)
This matches the target context provided (fission yeast, GH13 family, precursor/GPI-anchored cell surface protein) and avoids confusion with unrelated “aah1” symbols in other organisms. (morita2006anαamylasehomologue pages 3-5)
For the S. pombe paralog Aah3, catalytic-residue point mutations abolish normal function without altering localization, implying that enzymatic catalysis (not merely scaffolding) is required for its cell-wall role. However, soluble-starch hydrolysis assays on lysates/media showed no detectable starch-degrading activity, supporting the idea that Aah proteins are not conventional secreted starch hydrolases under tested conditions and may instead act as cell-wall transglycosylases on endogenous glucan substrates. (Morita et al., 2006; 2006-06; https://doi.org/10.1271/bbb.50693) (morita2006anαamylasehomologue pages 7-8)
Aah1 itself retains the conserved catalytic Asp/Glu/Asp triad and has predicted GPI anchoring, but Aah1-only biochemical assays and substrate panels were not identified in the evidence retrieved here. (morita2006anαamylasehomologue pages 3-5)
Two GPI-anchored Aspergillus niger GH13 enzymes (AgtA/AgtB) were biochemically shown to have 4-α-glucanotransferase activity (EC 2.4.1.25). Their reactions are dominated by transglycosylation: they cleave α-(1,4) bonds in donor substrates and form new α-(1,4) linkages in products. Key substrate rules include:
- Donor length requirement: donors require at least ~5 glucose units. (kaaij2007twonovelputatively pages 1-2, kaaij2007twonovelputatively pages 8-9)
- Acceptors: maltose is the smallest effective acceptor; larger maltooligosaccharides also act as acceptors. (kaaij2007twonovelputatively pages 1-2, kaaij2007twonovelputatively pages 8-9)
- Cross-linking potential: AgtA can accept small α-(1,3)-linked nigerooligosaccharides (e.g., nigerose/nigerotriose) as acceptors when combined with a (1,4)-linked donor, suggesting a possible biochemical bridge between α-1,4 and α-1,3 glucan contexts in cell walls. (kaaij2007twonovelputatively pages 8-9)
Because S. pombe Aah proteins cluster with these fungal GPI-anchored GH13 α-glucanotransferases in functional discussions and share the same domain logic (signal peptide + GPI anchor + α-amylase fold), these enzymological findings provide the best current, experimentally grounded substrate and reaction expectations for Aah1 when annotating function. (van der Kaaij et al., 2007; publication date: 2007-07; https://doi.org/10.1128/ec.00354-06) (kaaij2007twonovelputatively pages 1-2, kaaij2007twonovelputatively pages 5-6)
In pathogenic fungi, an α-(1,4)-amylase-like GH13 enzyme (Amy1) is genetically linked to cell-wall α-(1,3)-glucan production, and biochemical assays show it produces maltooligosaccharides of ~DP4–DP5 from starch/amylopectin. These products are proposed to serve as primer molecules for α-(1,3)-glucan synthase activity. (Camacho et al., 2012; publication date: 2012-11; https://doi.org/10.1371/journal.pone.0050201) (camacho2012expressionofparacoccidioides pages 1-2)
While this is not S. pombe Aah1, it supports a plausible mechanistic relationship between α-(1,4)-acting GH13 enzymes and the emergence/organization of α-(1,3)-glucan in fungal walls. (camacho2012expressionofparacoccidioides pages 1-2)
A recent S. pombe study using solid-state NMR and microscopy reports that aah1Δ aah3Δ double mutants are nearly spherical, grow slowly, show severe cell separation defects, and have thickened cell walls with altered outer layers. These data support that Aah1 (with Aah3) is required for normal cell-wall α-glucan architecture and mechanics. (Jacob et al., 2025 bioRxiv; publication date: 2025-03; https://doi.org/10.1101/2025.03.10.642465) (jacob2025solidstatenmranalysis pages 5-9, jacob2025solidstatenmranalysis pages 1-5)
Mechanistically, the authors interpret Aah1/Aah3 as cell-wall α-glucan remodeling enzymes acting alongside—but nonredundantly with—Ags1. They propose Aah1/Aah3 are likely downstream of Ags1 polymer synthesis to transglycosylate α-glucan chains into fibers of appropriate dimensions, which is required for proper rod shape and cytokinetic separation. (jacob2025solidstatenmranalysis pages 5-9, jacob2025solidstatenmranalysis pages 1-5)
In the 2006 study of the aah gene family, disruptions of aah1 were reported to be viable at 30 °C and 37 °C, indicating aah1 is not essential under those test conditions. In contrast, aah3 disruption showed clearer wall/morphogenesis phenotypes. (Morita et al., 2006; 2006-06; https://doi.org/10.1271/bbb.50693) (morita2006anαamylasehomologue pages 3-5)
The strong phenotype of the double mutant (aah1Δ aah3Δ) indicates substantial functional overlap or cooperation between Aah1 and Aah3 in maintaining α-glucan-rich wall architecture. (jacob2025solidstatenmranalysis pages 5-9, jacob2025solidstatenmranalysis pages 1-5)
Aah1 contains a C-terminal hydrophobic tail consistent with a GPI-anchor signal, supporting localization as a GPI-anchored membrane/cell-wall protein rather than a soluble cytosolic enzyme. (morita2006anαamylasehomologue pages 3-5)
Consistent with this class of proteins, the Aah-family enzymes are discussed as cell-surface/cell-wall–associated remodeling enzymes in the ssNMR study. (jacob2025solidstatenmranalysis pages 5-9, jacob2025solidstatenmranalysis pages 1-5)
The most information-dense quantitative dataset available here is the ssNMR/TEM analysis of wild type versus aah1Δ aah3Δ, which provides compositional shifts and wall-thickness changes.
| Genotype/condition | Key phenotype | Cell wall thickness (nm) | Rigid-phase composition (% α-1,3 glucan, % β-1,3 glucan) | Mobile-phase composition (total α-glucans %, total β-glucans %) | Notes on localization/biochemistry | Source (with DOI URL and year) |
|---|---|---|---|---|---|---|
| Wild type S. pombe | Rod-shaped cells; normal separation and cell wall architecture | ~100 | 44%, 54% | 33%, 48% | Reference condition for ssNMR/TEM comparisons (jacob2025solidstatenmranalysis media 9d9bf491, jacob2025solidstatenmranalysis media 9f92b336, jacob2025solidstatenmranalysis media b7bfd757) | Jacob et al., 2025, bioRxiv, https://doi.org/10.1101/2025.03.10.642465 |
| aah1Δ aah3Δ | Nearly spherical cells, slow growth, severe cell separation defects, clumping, thickened/abnormal walls, reduced FITC-ConA staining; viability depends on compensatory β-glucan upregulation/cell wall integrity pathway (jacob2025solidstatenmranalysis pages 9-12, jacob2025solidstatenmranalysis pages 5-9, jacob2025solidstatenmranalysis pages 1-5) | ~400 | 11%, 89% | 10%, 82% | Aah1/Aah3 described as putative GPI-anchored GH13 α-amylase-family cell-surface enzymes that likely act as α-glucan transglycosylases/remodelers rather than simple hydrolases; mutant loses major α-glucan matrix, including reduced α-1,3- and α-1,4-glucans (jacob2025solidstatenmranalysis pages 9-12, jacob2025solidstatenmranalysis pages 5-9, jacob2025solidstatenmranalysis pages 1-5, jacob2025solidstatenmranalysis pages 15-19) | Jacob et al., 2025, bioRxiv, https://doi.org/10.1101/2025.03.10.642465 |
| aah1Δ | No clear growth or morphology defect reported under tested conditions; nonessential for viability at 30 °C or 37 °C (morita2006anαamylasehomologue pages 3-5) | NR | NR | NR | Aah1 retains conserved GH13 catalytic Asp/Glu/Asp residues and has a C-terminal hydrophobic region consistent with a GPI-anchor signal; direct biochemical activity and substrate specificity for Aah1 alone not reported in this study (morita2006anαamylasehomologue pages 3-5) | Morita et al., 2006, Biosci. Biotechnol. Biochem., https://doi.org/10.1271/bbb.50693 |
| aah3Δ | Cell wall integrity and morphogenesis defects reported for single mutant in Morita et al.; among aah family disruptions, aah3Δ showed the clear phenotype (morita2006anαamylasehomologue pages 3-5) | NR | NR | NR | Aah3 characterized as a GPI-anchored membrane protein required for cell wall integrity and morphogenesis; used with Aah1 in later model of extracellular α-glucan remodeling (jacob2025solidstatenmranalysis pages 5-9, morita2006anαamylasehomologue pages 3-5) | Morita et al., 2006, Biosci. Biotechnol. Biochem., https://doi.org/10.1271/bbb.50693 |
| Aah1 protein prediction (sequence feature) | NR | NR | NR | NR | Predicted precursor/cell-surface protein with GPI-anchor signal; belongs to GH13/α-amylase family and preserves catalytic residues consistent with enzymatic activity on α-glucans, but no direct Aah1-only enzymology was shown in the cited experiments (morita2006anαamylasehomologue pages 3-5) | Morita et al., 2006, Biosci. Biotechnol. Biochem., https://doi.org/10.1271/bbb.50693 |
Table: This table compiles the most relevant phenotype, localization, and quantitative cell wall composition findings for S. pombe aah1/aah3 from the available evidence. It highlights that strong quantitative composition data are currently reported for the double mutant, while Aah1-only evidence is mainly sequence/localization-based.
Key quantitative findings include:
- Cell wall thickness: ~100 nm in wild type versus ~400 nm in aah1Δ aah3Δ. (jacob2025solidstatenmranalysis media 9d9bf491)
- Rigid-phase carbohydrate composition: α-1,3-glucan decreases from 44% to 11%, while β-1,3-glucan increases from 54% to 89% in aah1Δ aah3Δ. (jacob2025solidstatenmranalysis media 9f92b336)
- Mobile-phase composition: total α-glucans decrease from 33% to 10%, while total β-glucans increase from 48% to 82% in aah1Δ aah3Δ. (jacob2025solidstatenmranalysis media b7bfd757)
These shifts support an interpretation that Aah1/Aah3 are required to build or maintain the α-glucan matrix (both α-1,3 and minor α-1,4 components) and that the cell compensates by increasing β-glucan content via a cell wall integrity response. (jacob2025solidstatenmranalysis pages 1-5, jacob2025solidstatenmranalysis media 9f92b336, jacob2025solidstatenmranalysis media b7bfd757)
No 2023–2024 publications specifically interrogating S. pombe Aah1 were retrieved in this run. Nevertheless, 2023 research directly on EC 2.4.1.25 enzymes provides state-of-the-art mechanistic and engineering insights relevant to GH13/GH77 transglycosylases:
1) Domain fusion with starch-binding modules (CBM20) tunes substrate targeting and reaction partitioning. N-terminal fusion of starch-binding domains to a thermophilic 4-α-glucanotransferase increased activity on amylose 1.2–2.4×, increased hydrolytic activity 1.3–1.7×, and improved binding affinity to waxy maize starch granules (Kd 0.7–1.5 mg/mL for two fusions). (Wang et al., 2023; publication date: 2023-01; https://doi.org/10.3390/molecules28031320) (wang2023impactofstarch pages 1-2)
2) Fine structural elements in noncatalytic domains control hydrolysis vs transglycosylation. A domain-B loop in a bacterial 4-α-glucanotransferase acted as a “lid” limiting water access; W131G or loop deletion increased the hydrolysis/transglycosylation ratio 11.4–11.6× relative to wild type, demonstrating that modest structural edits can shift reaction specificity substantially. (Llopiz et al., 2023; publication date: 2023-07; https://doi.org/10.1007/s10930-023-10136-2) (llopiz2023theroleof pages 1-2)
Relevance to Aah1 annotation: these studies reinforce that α-amylase-fold enzymes can be biased toward transglycosylation by structural features, supporting the plausibility that GPI-anchored fungal GH13 proteins such as Aah1 operate primarily as cell-wall transglycosylases rather than starch hydrolases. (llopiz2023theroleof pages 1-2, wang2023impactofstarch pages 1-2)
4-α-glucanotransferases are actively engineered for starch modification, affecting molecular weight distributions and linkage patterns relevant to food and industrial starch functionality. For example, CBM20/SBD fusion engineering improved binding to granular starch and altered product linkage ratios in modified starch (α-1,6/α-1,4 ratio changes reported), illustrating a route to tailor starch properties using enzyme design. (wang2023impactofstarch pages 10-12, wang2023impactofstarch pages 1-2)
Because Aah1/Aah3-like proteins are cell-surface/cell-wall enzymes required for proper α-glucan matrix architecture in S. pombe, they represent a conceptual antifungal target class: extracellular remodeling enzymes acting in concert with synthases can be leveraged to disrupt wall mechanics. This inference is supported by the strong dependence of the aah1Δ aah3Δ mutant on compensatory β-glucan upregulation and the cell wall integrity pathway. (jacob2025solidstatenmranalysis pages 1-5)
Based on (i) GPI-anchoring and α-amylase catalytic-motif conservation, (ii) the severe α-glucan depletion and morphogenesis/separation defects in aah1Δ aah3Δ, and (iii) direct enzymology from close fungal GPI-anchored GH13 α-glucanotransferases, the most evidence-supported annotation for Aah1 is:
- Molecular function: α-glucan transglycosylase / 4-α-glucanotransferase-like activity acting on α-1,4-linked glucans (donors of at least ~DP5) and using maltose/maltooligosaccharides (and possibly α-1,3-linked oligosaccharides) as acceptors, producing remodeled/extended α-glucan chains. (kaaij2007twonovelputatively pages 1-2, kaaij2007twonovelputatively pages 8-9)
- Biological role: cell-wall α-glucan remodeling required for normal α-1,3/α-1,4 glucan matrix abundance and correct fiber dimensions, thereby supporting rod shape and cell separation. (jacob2025solidstatenmranalysis pages 5-9, jacob2025solidstatenmranalysis pages 1-5)
- Localization: secretory pathway → cell surface/cell wall via GPI anchoring. (morita2006anαamylasehomologue pages 3-5)
References
(jacob2025solidstatenmranalysis pages 9-12): Anand Jacob, Alaina H. Willet, Maya G. Igarashi, Mustapha El Hariri El Nokab, Lesley A. Turner, Abdulrahman Khalid A. Alsanad, Tuo Wang, and Kathleen L. Gould. Solid-state nmr analysis ofschizosaccharomyces pombereveals role ofα-amylase family enzymes in cell wall structure and function. BioRxiv, Mar 2025. URL: https://doi.org/10.1101/2025.03.10.642465, doi:10.1101/2025.03.10.642465. This article has 0 citations.
(jacob2025solidstatenmranalysis pages 5-9): Anand Jacob, Alaina H. Willet, Maya G. Igarashi, Mustapha El Hariri El Nokab, Lesley A. Turner, Abdulrahman Khalid A. Alsanad, Tuo Wang, and Kathleen L. Gould. Solid-state nmr analysis ofschizosaccharomyces pombereveals role ofα-amylase family enzymes in cell wall structure and function. BioRxiv, Mar 2025. URL: https://doi.org/10.1101/2025.03.10.642465, doi:10.1101/2025.03.10.642465. This article has 0 citations.
(jacob2025solidstatenmranalysis pages 1-5): Anand Jacob, Alaina H. Willet, Maya G. Igarashi, Mustapha El Hariri El Nokab, Lesley A. Turner, Abdulrahman Khalid A. Alsanad, Tuo Wang, and Kathleen L. Gould. Solid-state nmr analysis ofschizosaccharomyces pombereveals role ofα-amylase family enzymes in cell wall structure and function. BioRxiv, Mar 2025. URL: https://doi.org/10.1101/2025.03.10.642465, doi:10.1101/2025.03.10.642465. This article has 0 citations.
(vosUnknownyearlossofglucosidasea pages 45-48): A Vos, A de Haan, N Dekker, and F Hochstenbach. Loss of glucosidase ii results in-glucan accumulation in fission yeast. Unknown journal, Unknown year.
(kaaij2007twonovelputatively pages 1-2): R. M. van der Kaaij, X.-L. Yuan, A. Franken, A. F. J. Ram, P. J. Punt, M. J. E. C. van der Maarel, and L. Dijkhuizen. Two novel, putatively cell wall-associated and glycosylphosphatidylinositol-anchored α-glucanotransferase enzymes ofaspergillus niger. Jul 2007. URL: https://doi.org/10.1128/ec.00354-06, doi:10.1128/ec.00354-06. This article has 50 citations and is from a peer-reviewed journal.
(vosUnknownyearlossofglucosidase pages 183-189): A Vos, A de Haan, N Dekker, and F Hochstenbach. Loss of glucosidase ii results in-glucan accumulation in fission yeast. Unknown journal, Unknown year.
(vosUnknownyearevidenceforthea pages 183-189): A Vos, A de Haan, B Snijder, and F Hochstenbach. Evidence for the essential involvement of the transglycosylase domain of ags1p in cell wall-glucan biosynthesis in fission yeast. Unknown journal, Unknown year.
(morita2006anαamylasehomologue pages 3-5): Tomotake MORITA, Naotaka TANAKA, Akira HOSOMI, Yuko GIGA-HAMA, and Kaoru TAKEGAWA. An α-amylase homologue, aah3, encodes a gpi-anchored membrane protein required for cell wall integrity and morphogenesis in schizosaccharomyces pombe. Bioscience, Biotechnology, and Biochemistry, 70:1454-1463, Jun 2006. URL: https://doi.org/10.1271/bbb.50693, doi:10.1271/bbb.50693. This article has 31 citations.
(morita2006anαamylasehomologue pages 7-8): Tomotake MORITA, Naotaka TANAKA, Akira HOSOMI, Yuko GIGA-HAMA, and Kaoru TAKEGAWA. An α-amylase homologue, aah3, encodes a gpi-anchored membrane protein required for cell wall integrity and morphogenesis in schizosaccharomyces pombe. Bioscience, Biotechnology, and Biochemistry, 70:1454-1463, Jun 2006. URL: https://doi.org/10.1271/bbb.50693, doi:10.1271/bbb.50693. This article has 31 citations.
(kaaij2007twonovelputatively pages 8-9): R. M. van der Kaaij, X.-L. Yuan, A. Franken, A. F. J. Ram, P. J. Punt, M. J. E. C. van der Maarel, and L. Dijkhuizen. Two novel, putatively cell wall-associated and glycosylphosphatidylinositol-anchored α-glucanotransferase enzymes ofaspergillus niger. Jul 2007. URL: https://doi.org/10.1128/ec.00354-06, doi:10.1128/ec.00354-06. This article has 50 citations and is from a peer-reviewed journal.
(kaaij2007twonovelputatively pages 5-6): R. M. van der Kaaij, X.-L. Yuan, A. Franken, A. F. J. Ram, P. J. Punt, M. J. E. C. van der Maarel, and L. Dijkhuizen. Two novel, putatively cell wall-associated and glycosylphosphatidylinositol-anchored α-glucanotransferase enzymes ofaspergillus niger. Jul 2007. URL: https://doi.org/10.1128/ec.00354-06, doi:10.1128/ec.00354-06. This article has 50 citations and is from a peer-reviewed journal.
(camacho2012expressionofparacoccidioides pages 1-2): Emma Camacho, Victoria E. Sepulveda, William E. Goldman, Gioconda San-Blas, and Gustavo A. Niño-Vega. Expression of paracoccidioides brasiliensis amy1 in a histoplasma capsulatum amy1 mutant, relates an α-(1,4)-amylase to cell wall α-(1,3)-glucan synthesis. PLoS ONE, 7:e50201, Nov 2012. URL: https://doi.org/10.1371/journal.pone.0050201, doi:10.1371/journal.pone.0050201. This article has 36 citations and is from a peer-reviewed journal.
(jacob2025solidstatenmranalysis media 9d9bf491): Anand Jacob, Alaina H. Willet, Maya G. Igarashi, Mustapha El Hariri El Nokab, Lesley A. Turner, Abdulrahman Khalid A. Alsanad, Tuo Wang, and Kathleen L. Gould. Solid-state nmr analysis ofschizosaccharomyces pombereveals role ofα-amylase family enzymes in cell wall structure and function. BioRxiv, Mar 2025. URL: https://doi.org/10.1101/2025.03.10.642465, doi:10.1101/2025.03.10.642465. This article has 0 citations.
(jacob2025solidstatenmranalysis media 9f92b336): Anand Jacob, Alaina H. Willet, Maya G. Igarashi, Mustapha El Hariri El Nokab, Lesley A. Turner, Abdulrahman Khalid A. Alsanad, Tuo Wang, and Kathleen L. Gould. Solid-state nmr analysis ofschizosaccharomyces pombereveals role ofα-amylase family enzymes in cell wall structure and function. BioRxiv, Mar 2025. URL: https://doi.org/10.1101/2025.03.10.642465, doi:10.1101/2025.03.10.642465. This article has 0 citations.
(jacob2025solidstatenmranalysis media b7bfd757): Anand Jacob, Alaina H. Willet, Maya G. Igarashi, Mustapha El Hariri El Nokab, Lesley A. Turner, Abdulrahman Khalid A. Alsanad, Tuo Wang, and Kathleen L. Gould. Solid-state nmr analysis ofschizosaccharomyces pombereveals role ofα-amylase family enzymes in cell wall structure and function. BioRxiv, Mar 2025. URL: https://doi.org/10.1101/2025.03.10.642465, doi:10.1101/2025.03.10.642465. This article has 0 citations.
(jacob2025solidstatenmranalysis pages 15-19): Anand Jacob, Alaina H. Willet, Maya G. Igarashi, Mustapha El Hariri El Nokab, Lesley A. Turner, Abdulrahman Khalid A. Alsanad, Tuo Wang, and Kathleen L. Gould. Solid-state nmr analysis ofschizosaccharomyces pombereveals role ofα-amylase family enzymes in cell wall structure and function. BioRxiv, Mar 2025. URL: https://doi.org/10.1101/2025.03.10.642465, doi:10.1101/2025.03.10.642465. This article has 0 citations.
(wang2023impactofstarch pages 1-2): Yu Wang, Yazhen Wu, Stefan Jarl Christensen, Štefan Janeček, Yuxiang Bai, Marie Sofie Møller, and Birte Svensson. Impact of starch binding domain fusion on activities and starch product structure of 4-α-glucanotransferase. Molecules, 28:1320, Jan 2023. URL: https://doi.org/10.3390/molecules28031320, doi:10.3390/molecules28031320. This article has 11 citations.
(llopiz2023theroleof pages 1-2): Alexey Llopiz, Marco A. Ramírez-Martínez, Leticia Olvera, Wendy Xolalpa-Villanueva, Nina Pastor, and Gloria Saab-Rincon. The role of a loop in the non-catalytic domain b on the hydrolysis/transglycosylation specificity of the 4-α-glucanotransferase from thermotoga maritima. The Protein Journal, 42:502-518, Jul 2023. URL: https://doi.org/10.1007/s10930-023-10136-2, doi:10.1007/s10930-023-10136-2. This article has 3 citations.
(wang2023impactofstarch pages 10-12): Yu Wang, Yazhen Wu, Stefan Jarl Christensen, Štefan Janeček, Yuxiang Bai, Marie Sofie Møller, and Birte Svensson. Impact of starch binding domain fusion on activities and starch product structure of 4-α-glucanotransferase. Molecules, 28:1320, Jan 2023. URL: https://doi.org/10.3390/molecules28031320, doi:10.3390/molecules28031320. This article has 11 citations.
"α-glucan remodeling by GH13-domain enzymes shapes fungal cell wall architecture."
This is the primary experimental paper for aah1 function. Key findings:
- Aah1 and Aah3 are GPI-anchored GH13-family enzymes that act redundantly PMID:40668835
- Double mutant (aah1Δ aah3Δ) shows severe phenotype: rounded cells, delayed division, clumping PMID:40668835
- Solid-state NMR: α-1,3-glucan drops from 44% to 11% in rigid phase of double mutant PMID:40668835
- Cell wall thickens ~4x (100 nm to 400 nm) with compensatory β-glucan increase
- Proposed function: GH13-family transglycosylases that remodel α-glucan downstream of Ags1 synthase
"An alpha-amylase homologue, aah3, encodes a GPI-anchored membrane protein required for cell wall integrity and morphogenesis in S. pombe."
id: O74922
gene_symbol: aah1
product_type: PROTEIN
status: DRAFT
taxon:
id: NCBITaxon:284812
label: Schizosaccharomyces pombe (strain 972 / ATCC 24843)
description: >-
Aah1 is a GPI-anchored, cell-surface GH13-family enzyme that functions as an alpha-glucan
transglycosylase (4-alpha-glucanotransferase, EC 2.4.1.25) in S. pombe. It acts redundantly
with its paralog Aah3 to remodel alpha-glucan chains synthesized by the alpha-glucan synthase
Ags1, building a properly organized alpha-1,3-glucan matrix in the cell wall. Loss of both
aah1 and aah3 causes severe reduction of alpha-1,3-glucan, rounded cell morphology, delayed
cell division, clumping, and cell wall thickening with compensatory beta-glucan increases.
Single aah1 deletion has no obvious phenotype under standard conditions, indicating functional
overlap with aah3.
existing_annotations:
- term:
id: GO:0004134
label: 4-alpha-glucanotransferase activity
evidence_type: IEA
original_reference_id: GO_REF:0000003
review:
summary: >-
This IEA annotation from EC mapping to EC 2.4.1.25 is consistent with the UniProt-assigned
EC number and with the strong functional inference from close A. niger orthologs (AgtA/AgtB)
that are biochemically characterized as 4-alpha-glucanotransferases.
action: ACCEPT
reason: >-
The EC 2.4.1.25 assignment is well supported by domain conservation, catalytic triad
preservation, and direct enzymology of GPI-anchored GH13 fungal orthologs. The IEA
annotation is redundant with the ISS/ISO annotations below but consistent.
supported_by:
- reference_id: PMID:40668835
supporting_text: >-
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly
with the α-glucan synthase to build a properly organized α-glucan matrix.
- reference_id: file:SCHPO/aah1/aah1-deep-research-falcon.md
supporting_text: >-
Two GPI-anchored A. niger GH13 enzymes (AgtA/AgtB) were biochemically shown to have
4-alpha-glucanotransferase activity (EC 2.4.1.25), cleaving alpha-(1,4) bonds and forming
new alpha-(1,4) linkages.
- term:
id: GO:0005509
label: calcium ion binding
evidence_type: IEA
original_reference_id: GO_REF:0000002
review:
summary: >-
Calcium ion binding is inferred from the InterPro alpha-amylase C-terminal domain (IPR015340).
UniProt notes by similarity that Aah1 binds 2 calcium ions per subunit, based on conservation
with alpha-amylase family members. This is a structural/cofactor role common to GH13 enzymes.
action: KEEP_AS_NON_CORE
reason: >-
Calcium binding is a conserved structural feature of GH13 enzymes required for proper folding
and catalytic activity, but it is not the core function of Aah1. The annotation is likely
correct based on domain conservation but represents a cofactor requirement rather than the
primary molecular function.
supported_by:
- reference_id: file:SCHPO/aah1/aah1-uniprot.txt
supporting_text: >-
Binds 2 calcium ions per subunit. Calcium is inhibitory at high concentrations.
- term:
id: GO:0005886
label: plasma membrane
evidence_type: IEA
original_reference_id: GO_REF:0000044
review:
summary: >-
Aah1 is a GPI-anchored protein tethered to the plasma membrane on the extracellular
side. While technically attached to the plasma membrane, the more informative localization
is the external side of the plasma membrane / fungal-type cell wall where it carries out
its function.
action: KEEP_AS_NON_CORE
reason: >-
The plasma membrane annotation is correct but less informative than the external side of
plasma membrane (GO:0009897) annotation that is also present. The GPI anchor tethers the
protein to the membrane, but the functional localization is extracellular/cell wall.
supported_by:
- reference_id: PMID:40668835
supporting_text: >-
...two conserved glycosylphosphatidylinositol-anchored α-amylase-like enzymes,
Aah1 and Aah3, which act redundantly as key contributors to α-glucan network formation.
- reference_id: file:SCHPO/aah1/aah1-uniprot.txt
supporting_text: >-
Cell membrane; Lipid-anchor, GPI-anchor; Extracellular side.
- term:
id: GO:0005975
label: carbohydrate metabolic process
evidence_type: IEA
original_reference_id: GO_REF:0000002
review:
summary: >-
This is a very broad parent term for the more specific cell wall polysaccharide
biosynthetic processes that Aah1 is involved in. The annotation is technically correct
but overly general.
action: MARK_AS_OVER_ANNOTATED
reason: >-
The specific biological processes GO:0070600 (fungal-type cell wall (1->3)-alpha-glucan
biosynthetic process) and GO:0051278 (fungal-type cell wall polysaccharide biosynthetic
process) are more informative and already present. This broad parent term does not add
useful information.
- term:
id: GO:0009897
label: external side of plasma membrane
evidence_type: IEA
original_reference_id: GO_REF:0000117
review:
summary: >-
Aah1 is a GPI-anchored protein localized to the external side of the plasma membrane
where it acts on cell wall alpha-glucans. This is consistent with the GPI anchor
prediction and the functional role of remodeling extracellular alpha-glucan polymers.
action: ACCEPT
reason: >-
The external side of the plasma membrane is the correct and informative localization
for a GPI-anchored cell surface enzyme. Supported by the GPI-anchor signal in the
sequence and the known cell-wall-associated function.
supported_by:
- reference_id: PMID:40668835
supporting_text: >-
...two conserved glycosylphosphatidylinositol-anchored α-amylase-like enzymes,
Aah1 and Aah3, which act redundantly as key contributors to α-glucan network formation.
- reference_id: PMID:16751704
supporting_text: >-
...Aah3p is an N-glycosylated, GPI-anchored membrane protein localized in the membrane
and cell wall fractions.
- term:
id: GO:0016052
label: carbohydrate catabolic process
evidence_type: IEA
original_reference_id: GO_REF:0000002
review:
summary: >-
This annotation from InterPro IPR015340 (alpha-amylase C-terminal domain) suggests a
catabolic role. However, Aah1 functions as a transglycosylase rather than a hydrolase.
No starch-degrading activity was detected in S. pombe cell lysates or culture media.
The protein remodels cell wall alpha-glucans rather than degrading carbohydrates.
action: REMOVE
reason: >-
Aah1 is not involved in carbohydrate catabolism. It is a transglycosylase that remodels
alpha-glucan polymers in the cell wall. No hydrolytic/catabolic activity was detected
experimentally, and the protein's biological role is biosynthetic/remodeling rather
than degradative.
supported_by:
- reference_id: PMID:16751704
supporting_text: >-
Expression of aah3 alleles in which the conserved aspartic acid and glutamic acid
residues required for hydrolase activity were replaced with alanine residues failed
to rescue the morphological and ascal wall defects of aah3Delta cells.
- reference_id: PMID:40668835
supporting_text: >-
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate
nonredundantly with the α-glucan synthase to build a properly organized
α-glucan matrix.
- term:
id: GO:0004134
label: 4-alpha-glucanotransferase activity
evidence_type: ISS
original_reference_id: GO_REF:0000024
review:
summary: >-
ISS annotation based on manual transfer from ortholog A2QTS4 (A. niger AgtA). AgtA
was biochemically demonstrated to have 4-alpha-glucanotransferase activity on
alpha-(1,4)-linked glucan donors requiring at least ~5 glucose units, with maltose
as the smallest effective acceptor.
action: ACCEPT
reason: >-
This is the core molecular function of Aah1. The ISS transfer from the biochemically
characterized A. niger ortholog is well justified given the conserved GH13 catalytic
domain, GPI-anchor architecture, and the genetic evidence from PMID:40668835 supporting
a transglycosylase role in cell wall alpha-glucan remodeling.
supported_by:
- reference_id: PMID:40668835
supporting_text: >-
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly
with the α-glucan synthase to build a properly organized α-glucan matrix.
- reference_id: file:SCHPO/aah1/aah1-deep-research-falcon.md
supporting_text: >-
Two GPI-anchored A. niger GH13 enzymes (AgtA/AgtB) were biochemically shown to have
4-alpha-glucanotransferase activity. Their reactions are dominated by transglycosylation:
they cleave alpha-(1,4) bonds in donor substrates and form new alpha-(1,4) linkages
in products.
- term:
id: GO:0004134
label: 4-alpha-glucanotransferase activity
evidence_type: ISO
original_reference_id: GO_REF:0000024
review:
summary: >-
ISO annotation from PomBase, also based on orthology to A2QTS4. This is the same
functional assignment as the ISS annotation above but from a different source (PomBase
vs UniProt).
action: ACCEPT
reason: >-
Consistent with the ISS annotation and well supported by the same evidence. The ISO
evidence code reflects orthology-based transfer, which is appropriate given the high
domain conservation and functional similarity demonstrated by genetics.
supported_by:
- reference_id: PMID:40668835
supporting_text: >-
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly
with the α-glucan synthase to build a properly organized α-glucan matrix.
- term:
id: GO:0051278
label: fungal-type cell wall polysaccharide biosynthetic process
evidence_type: ISS
original_reference_id: GO_REF:0000024
review:
summary: >-
ISS annotation transferred from ortholog. Aah1 contributes to biosynthesis of
cell wall alpha-glucan polysaccharides. The aah1/aah3 double mutant shows dramatic
reduction in cell wall alpha-1,3-glucan (from 44% to 11% in rigid phase).
action: ACCEPT
reason: >-
Aah1 is directly involved in building the alpha-glucan component of the fungal cell
wall. While the single mutant has no clear phenotype, the double mutant with aah3
demonstrates that Aah1 redundantly contributes to cell wall polysaccharide biosynthesis.
supported_by:
- reference_id: PMID:40668835
supporting_text: >-
...the double mutant cell walls have dramatically reduced α-1,3-glucan and
galactomannan content, with a compensatory increase in β-glucans driven by the
activation of the cell integrity pathway.
- term:
id: GO:0071940
label: fungal-type cell wall assembly
evidence_type: ISS
original_reference_id: GO_REF:0000024
review:
summary: >-
ISS annotation for cell wall assembly. Aah1 contributes to proper cell wall
architecture by remodeling alpha-glucan chains into mature organized fibers.
The double mutant has severely altered wall structure with 4-fold thickening.
action: ACCEPT
reason: >-
Cell wall assembly is a core biological process for Aah1. The protein remodels
alpha-glucan polymers synthesized by Ags1 into a properly organized matrix, which
is essential for normal cell wall architecture, rod shape, and cell separation.
supported_by:
- reference_id: PMID:40668835
supporting_text: >-
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly
with the α-glucan synthase to build a properly organized α-glucan matrix...
Cells lacking both enzymes exhibit severe growth and morphological defects, including
rounded shape, delayed division, and cell clumping.
- term:
id: GO:0070600
label: fungal-type cell wall (1->3)-alpha-glucan biosynthetic process
evidence_type: IMP
original_reference_id: PMID:40668835
review:
summary: >-
IMP annotation from PomBase based on the Jacob et al. 2025 study. The aah1/aah3
double mutant shows dramatic reduction in alpha-1,3-glucan (from 44% to 11% in rigid
phase). This is the most specific and informative biological process annotation.
action: ACCEPT
reason: >-
This is the most specific and experimentally supported biological process annotation
for Aah1. The IMP evidence from solid-state NMR analysis directly demonstrates that
loss of Aah1 (with Aah3) leads to severe reduction in alpha-1,3-glucan in the cell
wall. This represents the core biosynthetic process in which Aah1 participates.
supported_by:
- reference_id: PMID:40668835
supporting_text: >-
...the double mutant cell walls have dramatically reduced α-1,3-glucan and
galactomannan content, with a compensatory increase in β-glucans driven by the
activation of the cell integrity pathway.
- reference_id: file:SCHPO/aah1/aah1-deep-research-falcon.md
supporting_text: >-
Rigid-phase carbohydrate composition: alpha-1,3-glucan decreases from 44% to 11%
in aah1/aah3 double mutant.
- term:
id: GO:0009277
label: fungal-type cell wall
evidence_type: ISS
original_reference_id: GO_REF:0000024
review:
summary: >-
ISS annotation from PomBase for localization to the fungal-type cell wall. Aah1 is a
GPI-anchored enzyme that localizes to the cell surface/cell wall where it remodels
alpha-glucan polymers. This localization is consistent with the GPI-anchor signal and
the known cell wall function.
action: ACCEPT
reason: >-
As a GPI-anchored cell surface enzyme that remodels cell wall alpha-glucans, localization
to the fungal-type cell wall is appropriate. The paralog Aah3 was shown to localize to
membrane and cell wall fractions biochemically.
supported_by:
- reference_id: PMID:16751704
supporting_text: >-
...Aah3p is an N-glycosylated, GPI-anchored membrane protein localized in the membrane
and cell wall fractions.
- reference_id: PMID:40668835
supporting_text: >-
...two conserved glycosylphosphatidylinositol-anchored α-amylase-like enzymes,
Aah1 and Aah3, which act redundantly as key contributors to α-glucan network formation.
references:
- id: GO_REF:0000002
title: Gene Ontology annotation through association of InterPro records with GO terms
findings:
- statement: >-
InterPro domain IPR015340 (alpha-amylase C-terminal domain) maps to calcium ion binding
and carbohydrate metabolic/catabolic process GO terms.
- id: GO_REF:0000003
title: Gene Ontology annotation based on Enzyme Commission mapping
findings:
- statement: >-
EC 2.4.1.25 maps to 4-alpha-glucanotransferase activity (GO:0004134).
- id: GO_REF:0000024
title: Manual transfer of experimentally-verified manual GO annotation data to orthologs
by curator judgment of sequence similarity
findings:
- statement: >-
Orthology-based transfer from A. niger AgtA (A2QTS4) for molecular function and from
other S. pombe cell wall proteins for localization and biological process annotations.
- id: GO_REF:0000044
title: Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location
vocabulary mapping, accompanied by conservative changes to GO terms applied by UniProt
findings:
- statement: >-
Maps UniProt subcellular location 'Cell membrane' to GO:0005886 (plasma membrane).
- id: GO_REF:0000117
title: Electronic Gene Ontology annotations created by ARBA machine learning models
findings:
- statement: >-
ARBA rule ARBA00043397 predicts external side of plasma membrane localization.
- id: PMID:40668835
title: α-glucan remodeling by GH13-domain enzymes shapes fungal cell wall architecture.
findings:
- statement: >-
Aah1 and Aah3 are GPI-anchored GH13-family transglycosylases that act redundantly
to remodel alpha-glucan chains synthesized by Ags1 into a properly organized matrix.
supporting_text: >-
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly
with the α-glucan synthase to build a properly organized α-glucan matrix.
- statement: >-
aah1/aah3 double mutant shows severe growth defects, rounded cell shape, delayed
division, cell clumping, and dramatically altered cell wall composition.
supporting_text: >-
Cells lacking both enzymes exhibit severe growth and morphological defects, including
rounded shape, delayed division, and cell clumping.
- statement: >-
Solid-state NMR shows alpha-1,3-glucan decreases from 44% to 11% in rigid phase
of double mutant, with compensatory beta-glucan increase from 54% to 89%.
supporting_text: >-
...the double mutant cell walls have dramatically reduced α-1,3-glucan and
galactomannan content, with a compensatory increase in β-glucans driven by the
activation of the cell integrity pathway.
- statement: >-
Cell wall thickness increases approximately 4-fold in the double mutant, with
increased rigidity and reduced polymer mobility.
supporting_text: >-
These changes correlate with cell wall thickening, increased rigidity, and reduced
polymer mobility and hydration.
- id: PMID:16751704
title: An alpha-amylase homologue, aah3, encodes a GPI-anchored membrane protein required
for cell wall integrity and morphogenesis in Schizosaccharomyces pombe.
findings:
- statement: >-
Identified four alpha-amylase homologs (aah1-aah4) in S. pombe with putative signal
sequences and C-terminal GPI anchor addition signals.
supporting_text: >-
...four alpha-amylase homologs (Aah1p-Aah4p) have putative signal sequences and
C-terminal GPI anchor addition signals.
- statement: >-
Aah3 is a GPI-anchored membrane protein localized in the membrane and cell wall fractions.
supporting_text: >-
Biochemical analysis showed that Aah3p is an N-glycosylated, GPI-anchored membrane protein
localized in the membrane and cell wall fractions.
- statement: >-
Catalytic-residue point mutations in Aah3 abolished function without altering
localization, confirming enzymatic activity is required for the cell wall role.
supporting_text: >-
Expression of aah3 alleles in which the conserved aspartic acid and glutamic acid residues
required for hydrolase activity were replaced with alanine residues failed to rescue the
morphological and ascal wall defects of aah3Delta cells.
core_functions:
- molecular_function:
id: GO:0004134
label: 4-alpha-glucanotransferase activity
description: >-
Aah1 is a GH13-family enzyme that catalyzes transglycosylation of alpha-glucan chains,
transferring segments of alpha-(1,4)-linked glucan to acceptor molecules. Based on
characterized fungal orthologs, it likely cleaves alpha-(1,4) bonds in donor substrates
of at least ~5 glucose units and transfers them to acceptors including maltose and
maltooligosaccharides, and possibly alpha-(1,3)-linked oligosaccharides.
directly_involved_in:
- id: GO:0070600
label: fungal-type cell wall (1->3)-alpha-glucan biosynthetic process
- id: GO:0051278
label: fungal-type cell wall polysaccharide biosynthetic process
- id: GO:0071940
label: fungal-type cell wall assembly
locations:
- id: GO:0009897
label: external side of plasma membrane
- id: GO:0009277
label: fungal-type cell wall
supported_by:
- reference_id: PMID:40668835
supporting_text: >-
...Aah1 and Aah3 function as GH13-family transglycosylases that collaborate nonredundantly
with the α-glucan synthase to build a properly organized α-glucan matrix.
- reference_id: PMID:16751704
supporting_text: >-
...Aah3p is an N-glycosylated, GPI-anchored membrane protein localized in the membrane
and cell wall fractions.
suggested_questions:
- question: >-
What is the direct enzymatic activity and substrate specificity of purified Aah1?
No in vitro assay on purified Aah1 alone has been reported.
- question: >-
Does Aah1 have any unique substrates or products compared to Aah3, given that
they are described as acting nonredundantly with each other?
- question: >-
What is the individual contribution of aah1 to cell wall alpha-glucan under
stress conditions or during specific cell cycle stages?
suggested_experiments:
- description: >-
Purify recombinant Aah1 and characterize its substrate specificity and products
using defined alpha-glucan oligosaccharides as donors and acceptors.
- description: >-
Perform solid-state NMR analysis on aah1 single mutant (comparing to wild type and
aah3 single mutant) to determine the individual contribution of Aah1 to cell wall
composition.