LPXK (At3g20480) is a nuclear-encoded, mitochondrially targeted tetraacyldisaccharide 4'-kinase of the LpxK family, the single Arabidopsis ortholog of E. coli LpxK. By orthology it uses ATP to phosphorylate the 4'-position of the lipid A disaccharide 1-phosphate (DS-1-P), forming lipid IVA. This is the sixth step of a plant lipid A precursor pathway that mirrors the E. coli Raetz pathway (LpxA, LpxC, LpxD, LpxB, LpxK, KdtA). Three independent lpxk null alleles accumulate DS-1-P, which is undetectable in wild type, but the plants have no visible phenotype. An N-terminal GFP fusion is targeted to mitochondria. The final plant lipid A-like product and its physiological role, possibly a mitochondrial membrane component or a signalling lipid, are unknown.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005524 ATP binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: ATP binding from the InterPro2GO mapping of IPR003758 (LpxK family). Reason: LpxK is an ATP-dependent kinase with a P-loop nucleotide-binding fold, so ATP binding is correct. It is a sub-function of the core lipid-A 4'-kinase activity rather than an informative function in its own right. ATP binding is not an is_a ancestor of the kinase term, so it is kept as non-core and not marked too general. Supporting Evidence: file:ARATH/LPXK/LPXK-uniprot.txt Belongs to the LpxK family. PMID:21709257 In E. coli , LpxK phosphorylates the disaccharide 1-phosphate product made by LpxB to generate lipid IV A ( Fig. 1 ) ( 1 , 2 ). |
| GO:0005739 mitochondrion | IDA PMID:21709257 Pathway for lipid A biosynthesis in Arabidopsis thaliana res... | ACCEPT | Summary: Mitochondrion by GFP fusion and MitoTracker co-localisation in transgenic Arabidopsis roots (Li et al. 2011). Reason: The full-length AtLpxK-GFP fusion was too dim, but the 5'-terminal fusion showed the same mitochondrial pattern as AtLpxA and AtLpxC, and in mesophyll the Lpx fusion dots did not co-localise with chloroplasts. This agrees with the predicted 32-residue mitochondrial transit peptide. The sub-mitochondrial location is unknown, so mitochondrion is the right precision. Supporting Evidence: PMID:21709257 However, 5β²-terminal fusions of AtLpxD1, AtLpxD2, AtLpxB, and AtLpxK to GFP showed the same mitochondrial pattern as full-length AtLpxA and AtLpxC PMID:21709257 In mesophyll cells, GFP signals were shown as 1 ΞΌm dots, but were not colocalized with chloroplasts (data not shown). |
| GO:0005739 mitochondrion | IEA GO_REF:0000044 | ACCEPT | Summary: Mitochondrion from the UniProt subcellular-location mapping, which rests on Li et al. 2011 (ECO:0000269). Reason: Same claim as the IDA row; backed by the GFP fusion experiment. Supporting Evidence: file:ARATH/LPXK/LPXK-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion {ECO:0000269|PubMed:21709257}. PMID:21709257 However, 5β²-terminal fusions of AtLpxD1, AtLpxD2, AtLpxB, and AtLpxK to GFP showed the same mitochondrial pattern as full-length AtLpxA and AtLpxC |
| GO:0009029 lipid-A 4'-kinase activity | IBA GO_REF:0000033 | ACCEPT | Summary: Lipid-A 4'-kinase activity propagated from PAINT node PTN002014367, seeded by E. coli LpxK (P27300) and AtLpxK itself. Reason: AtLpxK is a full-length single-copy LpxK family member (26% identity to E. coli LpxK) inside the clade that inherited the activity. Its own IMP evidence (accumulation of the LpxK substrate DS-1-P in lpxk nulls) is part of the grounding, which is why AT3G20480 is in the WITH/FROM. Core function. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN002014367 SUPPORTS TRANSFER LpxK ancestral node; AtLpxK is a single-copy ortholog with no evidence of divergence. UniProtKB:P27300 SUPPORTS TRANSFER Biochemically characterised E. coli tetraacyldisaccharide 4'-kinase. Supporting Evidence: PMID:21709257 Single copy genes encoding AtLpxB (AT2G04560), AtLpxK (AT3G20480) and AtKdtA (AT5G03770) ( Table 1 ) were also found, with 31, 26, and 34% sequence identity to the corresponding E. coli proteins, respectively PMID:21709257 The disaccharide 1-phosphate was not seen in wild-type Arabidopsis , but accumulates in the atlpxk-1 and the atkdta-1 mutants |
| GO:0009029 lipid-A 4'-kinase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Lipid-A 4'-kinase activity from combined InterPro (IPR003758), Rhea and EC 2.7.1.130 mappings. Reason: Same core claim as the IBA and IMP rows. Supporting Evidence: file:ARATH/LPXK/LPXK-uniprot.txt Belongs to the LpxK family. PMID:21709257 The disaccharide 1-phosphate was not seen in wild-type Arabidopsis , but accumulates in the atlpxk-1 and the atkdta-1 mutants |
| GO:0009029 lipid-A 4'-kinase activity | IMP PMID:21709257 Pathway for lipid A biosynthesis in Arabidopsis thaliana res... | ACCEPT | Summary: Lipid-A 4'-kinase activity by IMP. Three independent lpxk null alleles accumulate the LpxK substrate, disaccharide 1-phosphate, which is undetectable in wild type. Reason: Loss of AtLpxK causes accumulation of DS-1-P, the substrate of the LpxK reaction. Its product, lipid IVA, is seen only when the next enzyme (KdtA) is missing. This is strong genetic evidence for the 4'-kinase step. No in vitro assay has been reported, but together with orthology the IMP is sound. Core function. Supporting Evidence: PMID:21709257 The disaccharide 1-phosphate was not seen in wild-type Arabidopsis , but accumulates in the atlpxk-1 and the atkdta-1 mutants PMID:21709257 The independently isolated atlpxb-2 , atlpxk-2 , and atlpxk-3 mutants showed similar lipid accumulation patterns as the atlpxb-1 and atlpxk-1 mutants. PMID:21709257 In E. coli , LpxK phosphorylates the disaccharide 1-phosphate product made by LpxB to generate lipid IV A ( Fig. 1 ) ( 1 , 2 ). |
| GO:0009245 lipid A biosynthetic process | IEA GO_REF:0000002 | ACCEPT | Summary: Lipid A biosynthetic process from the InterPro2GO mapping of IPR003758 (LpxK). This is part of the plant lipid A set flagged by Wei et al. 2020 (RAR), and the flag is refuted. Reason: AtLpxK catalyses the 4'-phosphorylation step of the lipid A precursor pathway, so it does the work of the process. In planta, lpxk nulls accumulate its substrate DS-1-P, and lipid X rises 2.6-fold, as expected for a block downstream of LpxB. The UniProt pathway line (step 6/6 of lipid IV(A) biosynthesis) rests on this experiment. This row also carries the experimental (IMP) evidence, which in GOA is attached only to the obsolete lipid X term. Supporting Evidence: PMID:21709257 our work demonstrates that plants synthesize lipid A precursors using the same enzymatic pathway present in E. coli PMID:21709257 The disaccharide 1-phosphate was not seen in wild-type Arabidopsis , but accumulates in the atlpxk-1 and the atkdta-1 mutants file:ARATH/LPXK/LPXK-uniprot.txt acetyl-alpha-D-glucosamine: step 6/6. {ECO:0000269|PubMed:21709257}. |
| GO:2001289 obsolete lipid X metabolic process | IMP PMID:21709257 Pathway for lipid A biosynthesis in Arabidopsis thaliana res... | MODIFY | Summary: Obsolete lipid X metabolic process, based on the 2.6-fold rise of lipid X in atlpxk-1. Reason: GO:2001289 is obsolete (QuickGO lists it as made in error and an unnecessary grouping term, with no replacement). LpxK does not act on lipid X. The modest lipid X increase is an indirect effect of blocking the pathway two steps downstream. The direct finding, accumulation of the LpxK substrate DS-1-P, supports lipid A biosynthetic process, which the gene already carries (IEA). Proposed replacements: lipid A biosynthetic process Supporting Evidence: PMID:21709257 In the atlpxk-1 and atkdta-1 mutants, lipid X levels were 2.6- and 2.2-fold higher than in the Col-0 wild type, respectively PMID:21709257 The disaccharide 1-phosphate was not seen in wild-type Arabidopsis , but accumulates in the atlpxk-1 and the atkdta-1 mutants |
Loading supporting contentβ¦
Download this section (compressed HTML)Q: Does recombinant mature AtLpxK phosphorylate DS-1-P in vitro, and does it complement an E. coli lpxK mutant despite its low (26%) identity?
Q: Is AtLpxK an integral membrane protein of the mitochondrial inner membrane, as E. coli LpxK is anchored in the inner membrane by an N-terminal helix?
Q: Which of the several EST-supported splice isoforms of At3g20480 encode an active, mitochondrially targeted kinase?
Experiment: Express mature AtLpxK in E. coli membranes, assay DS-1-P phosphorylation with [gamma-32P]ATP by TLC or LC-ESI/MS, and test rescue of an E. coli lpxK depletion strain.
Hypothesis: AtLpxK is a functional ATP-dependent tetraacyldisaccharide 4'-kinase.
Type: in vitro enzyme assay and heterologous complementation
Experiment: Carbonate extraction and protease protection of mitochondria from lines expressing full-length tagged LPXK under its native promoter.
Hypothesis: AtLpxK is anchored in a mitochondrial membrane.
Type: sub-mitochondrial fractionation
Loading supporting contentβ¦
Download this section (compressed HTML)Loading supporting contentβ¦
Download this section (compressed HTML)