TIM9 is a small mitochondrial intermembrane space (IMS) chaperone that forms a hexameric complex with TIM10 (the Tim9-Tim10 or TIM10 complex, composed of 3 copies of each subunit). This soluble 70 kDa complex functions as a carrier-holdase that escorts hydrophobic transmembrane protein precursors, especially mitochondrial carriers, from the TOM complex across the aqueous IMS to the TIM22 complex for insertion into the inner membrane. Small Tim complexes have also been implicated in delivery of some beta-barrel precursors toward the SAM complex. TIM9 contains a twin CX3C motif that forms two intramolecular disulfide bonds in the IMS; during cytoplasmic transit these cysteines may coordinate zinc. TIM9 is essential for viability and plays both a structural role in complex assembly and a functional role in substrate recognition, with its N-terminal region required for efficient trapping of incoming substrates.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005743 mitochondrial inner membrane | IBA GO_REF:0000033 | ACCEPT | Summary: TIM9 is a peripheral membrane protein on the IMS face of the mitochondrial inner membrane (PMID:9822593, PMID:9889188). The IBA annotation to mitochondrial inner membrane is phylogenetically consistent and supported by experimental evidence in yeast. UniProt records the subcellular location as "Mitochondrion inner membrane; Peripheral membrane protein; Intermembrane side." Reason: TIM9 is well-established as associated with the mitochondrial inner membrane, where a fraction of Tim9 is part of the membrane-associated 300 kDa TIM22 complex (PMID:9822593). The IBA annotation is phylogenetically sound and consistent with the IDA annotations. Propagation Review Root cause: SOURCE STALE OR MISSING Sources checked: PANTHER:PTN004407763 · PANTHER:PTN004407763 SOURCE STALE OR MISSING GOA retains this location IBA, but current PAINT no longer carries GO:0005743 at PTN004407763; the node now carries GO:0042719 complex membership. SGD:S000007256 · TIM9 SUPPORTS TRANSFER Target experimental evidence validly grounded the earlier ancestral call. UniProtKB:Q9Y5J6 · TIMM10B SUPPORTS TRANSFER Human TIMM10B is a conserved small-Tim family component. UniProtKB:Q9Y5J7 · TIMM9 SUPPORTS TRANSFER Human TIMM9 is the direct conserved ortholog. Supporting Evidence: PMID:9822593 A small fraction of Tim9p is bound to the outer face of the inner membrane in a 300 kDa complex whose other subunits include Tim54p, Tim22p, Tim12p and Tim10p. PMID:9889188 Tim9 is located in the mitochondrial intermembrane space and is organized into two distinct hetero-oligomeric assemblies with Tim10 and Tim12. |
| GO:0045039 protein insertion into mitochondrial inner membrane | IBA GO_REF:0000033 | ACCEPT | Summary: The Tim9-Tim10 complex escorts carrier protein precursors across the IMS to the TIM22 complex for insertion into the inner membrane. This is a core function of TIM9, supported by multiple experimental studies (PMID:9822593, PMID:9889188, PMID:10469659, PMID:11483513). The IBA annotation is phylogenetically consistent. Reason: This is the core biological process that TIM9 participates in. The Tim9-Tim10 complex mediates partial translocation of mitochondrial carrier proteins across the outer membrane and their subsequent insertion into the inner membrane via TIM22 (PMID:9889188). Multiple experimental evidence codes support this for yeast TIM9 directly. Propagation Review Root cause: SOURCE STALE OR MISSING Sources checked: PANTHER:PTN004407763 · PANTHER:PTN004407763 SOURCE STALE OR MISSING GOA retains this process IBA, but current PAINT no longer carries GO:0045039 at PTN004407763; the target's direct evidence independently supports the process. SGD:S000007256 · TIM9 SUPPORTS TRANSFER Target experimental evidence directly supports the insertion pathway role. MGI:MGI:1353436 · MGI:MGI:1353436 SUPPORTS TRANSFER Mammalian ortholog evidence supports conservation of the pathway role. UniProtKB:Q9Y5J7 · TIMM9 SUPPORTS TRANSFER Human TIMM9 supports the conserved small-Tim pathway assignment. WB:WBGene00006572 · WB:WBGene00006572 SUPPORTS TRANSFER Nematode ortholog evidence supports the conserved process. Supporting Evidence: PMID:9889188 The TIM9.10 complex is more abundant than the TIM9.10.12 complex and mediates partial translocation of mitochondrial carriers proteins across the outer membrane. PMID:9822593 Tim9p is a new subunit of the TIM machinery that guides hydrophobic inner membrane proteins across the aqueous intermembrane space. file:yeast/TIM9/TIM9-deep-research-falcon.md Tim9 acts as a **chaperone/escort factor** that prevents aggregation of hydrophobic precursors in the aqueous IMS and transfers them from **TOM** to **TIM22**. |
| GO:0140318 protein transporter activity | IBA GO_REF:0000033 | ACCEPT | Summary: TIM9 functions in the Tim9-Tim10 shuttle that directly binds hydrophobic precursors and delivers them through the IMS. Live GO explicitly cites the soluble Tim9-Tim10 complex as an example of GO:0140318 protein transporter activity. GO:0140309 unfolded protein holdase activity adds the substrate-state and anti-aggregation dimension, but does not invalidate this transporter annotation. Reason: GO:0140318 precisely captures TIM9's directed binding and delivery of precursor proteins between TOM-side intermediates and downstream insertases. It is retained as a core activity alongside the more specific holdase annotation. Propagation Review Root cause: SOURCE STALE OR MISSING Sources checked: PANTHER:PTN004407763 · PANTHER:PTN004407763 SOURCE STALE OR MISSING GOA retains this MF IBA, but current PAINT no longer carries GO:0140318 at PTN004407763; direct target IDA and IGI rows independently support the activity. SGD:S000007256 · TIM9 SUPPORTS TRANSFER The target's own experimental annotation validly grounded the ancestral call; target-in-own-WITH/FROM is not circular. Supporting Evidence: PMID:9822593 Tim9p is a new subunit of the TIM machinery that guides hydrophobic inner membrane proteins across the aqueous intermembrane space. |
| GO:0005743 mitochondrial inner membrane | IEA GO_REF:0000044 | ACCEPT | Summary: IEA annotation mapping from UniProtKB subcellular location. TIM9 is annotated in UniProt as "Mitochondrion inner membrane; Peripheral membrane protein; Intermembrane side." This is consistent with experimental evidence. Reason: This IEA is broader than the IBA and IDA annotations for the same term and is consistent with the known localization of TIM9 at the mitochondrial inner membrane (PMID:9822593). Supporting Evidence: UniProt:O74700 SUBCELLULAR LOCATION: Mitochondrion inner membrane |
| GO:0005758 mitochondrial intermembrane space | IEA GO_REF:0000044 | ACCEPT | Summary: IEA annotation mapping from UniProtKB subcellular location. TIM9 is localized to the mitochondrial intermembrane space where it forms the soluble Tim9-Tim10 70 kDa complex. Well supported by experimental evidence. Reason: The IMS localization is well-established experimentally (PMID:9822593, PMID:9889188) and confirmed by proteomics (PMID:22984289, cited in UniProt). The IEA is consistent with direct evidence. Supporting Evidence: PMID:9889188 Tim9 is located in the mitochondrial intermembrane space and is organized into two distinct hetero-oligomeric assemblies with Tim10 and Tim12. file:yeast/TIM9/TIM9-deep-research-falcon.md Tim9 is located in the **mitochondrial IMS** |
| GO:0015031 protein transport | IEA GO_REF:0000043 | KEEP AS NON CORE | Summary: IEA annotation from UniProtKB keyword mapping (KW-0653 Protein transport, KW-0811 Translocation). TIM9 participates in protein transport across the IMS. This is a general parent term; the more specific GO:0045039 (protein insertion into mitochondrial inner membrane) is also annotated. Reason: This umbrella is correct, but it is less informative than the reviewed transporter, holdase, and inner-membrane-insertion terms. It also accommodates the less directly documented SAM-directed route, so it is retained as non-core rather than narrowed to only one destination. Supporting Evidence: UniProt:O74700 Mitochondrial intermembrane chaperone that participates in the import and insertion of multi-pass transmembrane proteins into the mitochondrial inner membrane. |
| GO:0042719 mitochondrial intermembrane space chaperone complex | IEA GO_REF:0000117 | ACCEPT | Summary: IEA annotation from ARBA machine learning. TIM9 is a component of the mitochondrial intermembrane space chaperone complex (the Tim9-Tim10 hexameric complex). Well supported by experimental evidence. Reason: This IEA is consistent with the IDA annotations for the same term (PMID:9822593, PMID:9889188). TIM9 is a canonical subunit of the IMS chaperone complex. Falcon deep research independently confirms the Tim9-Tim10 heterohexamer with 3:3 stoichiometry (~70 kDa soluble complex). Supporting Evidence: PMID:9822593 Most of Tim9p is associated with Tim10p in a soluble 70 kDa complex. file:yeast/TIM9/TIM9-deep-research-falcon.md Tim9 forms the canonical **Tim9–Tim10 heterohexamer** and also participates in a **Tim9–Tim10–Tim12 docking complex** linked to TIM22. file:yeast/TIM9/TIM9-deep-research-falcon.md Tim9:Tim10 complex stoichiometry is **3:3**; the TIM22-associated complex is **Tim9–Tim10–Tim12 = 3:2:1**; the soluble Tim9/Tim10 complex is about **~70 kDa** |
| GO:0046872 metal ion binding | IEA GO_REF:0000043 | MARK AS OVER ANNOTATED | Summary: IEA annotation from UniProtKB keyword mapping (KW-0479 Metal-binding). TIM9 has a twin CX3C motif with four conserved cysteines. However, in the mature IMS-localized form, these cysteines form disulfide bonds rather than coordinating metal ions. Zinc coordination is thought to occur only transiently during cytoplasmic transit, and the mature functional form does not bind zinc (PMID:11867522). This IEA is misleading for the functional state of TIM9. Reason: While TIM9 may transiently coordinate zinc during its own import through the TOM complex, the mature functional form in the IMS contains disulfide bonds rather than zinc-coordinated cysteines (PMID:11867522). The UniProt entry itself states that the twin CX3C motif "contains 4 conserved Cys residues that form 2 disulfide bonds in the mitochondrial intermembrane space" and zinc coordination is only probable during transit. Annotating TIM9 as a metal ion binding protein is misleading for its functional state. Supporting Evidence: UniProt:O74700 The twin CX3C motif contains 4 conserved Cys residues that form 2 disulfide bonds in the mitochondrial intermembrane space. However, during the transit of TIM9 from cytoplasm into mitochondrion, the Cys residues probably coordinate zinc, thereby preventing folding and allowing its transfer across mitochondrial outer membrane file:yeast/TIM9/TIM9-deep-research-falcon.md Small TIM proteins (including Tim9) are imported through TOM and undergo oxidative folding in the IMS via the **MIA (mitochondrial import and assembly) pathway**, which introduces disulfides in the conserved CX3C motifs |
| GO:0005515 protein binding | IPI PMID:11483513 Functional reconstitution of the import of the yeast ADP/ATP... | MARK AS OVER ANNOTATED | Summary: IPI annotation based on physical interaction with TIM10 (PMID:11483513). Tim9 and Tim10 purified from E. coli can form a complex of the same size as the endogenous complex. This demonstrates direct protein-protein interaction. However, "protein binding" is uninformative -- TIM9 binding to TIM10 reflects its role in forming the functional hexameric chaperone complex. Reason: GO:0005515 "protein binding" is too vague and does not capture the specific functional interaction. TIM9 interacts with TIM10 to form the functional hexameric chaperone complex, and also binds to substrate carrier proteins during transit. The specific functions (unfolded protein holdase activity, complex membership) are captured by other annotations. Per curation guidelines, "protein binding" should be avoided in favor of more informative MF terms. Falcon deep research notes that within the complex Tim10 provides the key substrate-binding elements (Tim9 alone does not bind the AAC carrier), and that the obligate Tim9-Tim10 partnership forms an essential heterohexamer -- the functionally meaningful interaction, not generic protein binding. Supporting Evidence: PMID:11483513 Tim9 and Tim10 purified from Escherichia coli can form a complex of the same size as the endogenous complex from yeast mitochondria. This shows that no other mitochondrial protein is required for the formation of the TIM10 complex. file:yeast/TIM9/TIM9-deep-research-falcon.md Tim9 and Tim10 assemble into an essential heterohexameric Tim9/10 complex** that chaperones hydrophobic preproteins from TOM to TIM22 file:yeast/TIM9/TIM9-deep-research-falcon.md Biochemical substrate-binding evidence indicates that **Tim10** (alone) can bind the **ATP/ADP carrier (AAC)** similarly to the Tim9–Tim10 complex, whereas **Tim9 alone does not bind AAC**, supporting a model where Tim10 provides key substrate-binding elements within the complex |
| GO:0005739 mitochondrion | IDA PMID:9889188 Tim9, a new component of the TIM22.54 translocase in mitocho... | ACCEPT | Summary: IDA annotation for mitochondrial localization of TIM9 based on Adam et al. (1999), who identified Tim9 as a new component of the TIM22.54 translocase and demonstrated its mitochondrial localization. Reason: TIM9 is a well-established mitochondrial protein. This IDA is correct and supported by direct experimental evidence. Supporting Evidence: PMID:9889188 Tim9 is located in the mitochondrial intermembrane space and is organized into two distinct hetero-oligomeric assemblies with Tim10 and Tim12. |
| GO:0005743 mitochondrial inner membrane | IDA PMID:10648604 Tim18p, a new subunit of the TIM22 complex that mediates ins... | ACCEPT | Summary: IDA annotation for mitochondrial inner membrane localization based on Koehler et al. (2000), who characterized the TIM22 complex in the inner membrane containing Tim9p among its peripheral subunits. Reason: A fraction of TIM9 is associated with the inner membrane as part of the 300 kDa TIM22 complex. This is consistent with the UniProt annotation and the known biology of TIM9. Supporting Evidence: PMID:10648604 The TIM22 complex contains the peripheral subunits Tim9p, Tim10p, and Tim12p and the integral membrane subunits Tim22p and Tim54p. |
| GO:0005758 mitochondrial intermembrane space | IDA PMID:19037698 The Dutch multicenter experience of the endo-sponge treatmen... | ACCEPT | Summary: IDA annotation for IMS localization. However, PMID:19037698 is about "The Dutch multicenter experience of the endo-sponge treatment for anastomotic leakage after colorectal surgery" -- this is clearly an incorrect PMID. The intended reference is likely PMID:19037098 (Baker et al. 2009, about Tim9-Tim10 complex structure and function). The annotation itself is correct -- TIM9 is localized to the IMS -- but the reference is wrong. Reason: The localization of TIM9 to the mitochondrial IMS is well-established by multiple independent studies (PMID:9822593, PMID:9889188). Although the cited PMID:19037698 appears to be a data entry error (likely meant PMID:19037098), the annotation itself is correct. Note: the PMID should be corrected from 19037698 to 19037098. Supporting Evidence: PMID:9822593 Most of Tim9p is associated with Tim10p in a soluble 70 kDa complex. PMID:19037098 The Tim9-Tim10 complex plays an essential role in mitochondrial protein import by chaperoning select hydrophobic precursor proteins across the intermembrane space. |
| GO:0045039 protein insertion into mitochondrial inner membrane | IDA PMID:10648604 Tim18p, a new subunit of the TIM22 complex that mediates ins... | ACCEPT | Summary: IDA annotation based on Koehler et al. (2000), who showed that Tim9p is part of the TIM22 complex mediating insertion of carrier proteins into the inner membrane. Deletion of TIM18 (a TIM22 complex subunit) is synthetically lethal with temperature-sensitive mutations in Tim9p. Reason: This is a core function of TIM9. The Tim9-Tim10 complex escorts carrier proteins to the TIM22 complex for insertion into the inner membrane. Supporting Evidence: PMID:10648604 Deletion of Tim18p decreases the growth rate of yeast cells by a factor of two and is synthetically lethal with temperature-sensitive mutations in Tim9p or Tim10p. |
| GO:0045039 protein insertion into mitochondrial inner membrane | IDA PMID:19037098 Structural and functional requirements for activity of the T... | ACCEPT | Summary: IDA annotation based on Baker et al. (2009), who solved the Tim9-Tim10 crystal structure at 2.5 A resolution and performed mutational analysis showing that Tim9 plays an important functional role in facilitating the initial steps of translocating precursor substrates into the IMS. Reason: Baker et al. provide structural and functional evidence that Tim9 is directly involved in substrate recognition and translocation, not just complex stabilization. Supporting Evidence: PMID:19037098 We conclude that Tim9 plays an important functional role that includes facilitating the initial steps in translocating precursor substrates into the intermembrane space. |
| GO:0008270 zinc ion binding | RCA PMID:30358795 The cellular economy of the Saccharomyces cerevisiae zinc pr... | MARK AS OVER ANNOTATED | Summary: RCA annotation for zinc ion binding from Wang et al. (2018), a study characterizing the yeast zinc proteome by bioinformatic domain and motif searches. TIM9 was identified as a potential zinc-binding protein based on its CX3C motif. However, experimental evidence from Curran et al. (2002, PMID:11867522) demonstrated that TIM9 in the mature IMS form contains disulfide bonds, not zinc-coordinated cysteines. Reason: The RCA annotation is based on computational prediction from the CX3C motif. While TIM9 may transiently bind zinc during its cytoplasmic import phase, the mature functional form in the IMS uses disulfide bonds rather than zinc coordination (PMID:11867522, cited in UniProt). The annotation to GO:0008270 is misleading for the steady-state functional protein. Supporting Evidence: UniProt:O74700 The twin CX3C motif contains 4 conserved Cys residues that form 2 disulfide bonds in the mitochondrial intermembrane space. However, during the transit of TIM9 from cytoplasm into mitochondrion, the Cys residues probably coordinate zinc PMID:30358795 The yeast zinc proteome of 582 known or potential zinc-binding proteins was identified using a bioinformatics analysis that combined global domain searches with local motif searches. file:yeast/TIM9/TIM9-deep-research-falcon.md the conserved **CX3C** cysteines form **two intramolecular disulfide bonds** important for structural integrity |
| GO:0005758 mitochondrial intermembrane space | TAS Reactome:R-SCE-1252259 | ACCEPT | Summary: TAS annotation from Reactome pathway "TIM9:TIM10 binds hydrophobic proteins." The IMS localization is consistent with all experimental evidence. Reason: Correctly reflects the localization of TIM9 in the IMS as part of the Tim9-Tim10 complex. Redundant with IDA annotations but not incorrect. Supporting Evidence: Reactome:R-SCE-1252259 TIM9:TIM10 binds hydrophobic proteins |
| GO:0005758 mitochondrial intermembrane space | TAS Reactome:R-SCE-1252260 | ACCEPT | Summary: TAS annotation from Reactome pathway "MIA40:ERV1 oxidizes cysteine residues to cystine disulfide bonds." This Reactome entry relates to the MIA pathway by which TIM9 is imported and oxidized in the IMS. The IMS localization is correct. Reason: The MIA pathway (Mia40/Erv1) is responsible for the oxidative folding of TIM9 in the IMS, forming the disulfide bonds in its twin CX3C motif. This correctly places TIM9 in the IMS. Redundant with other IMS localization annotations but not incorrect. Supporting Evidence: Reactome:R-SCE-1252260 MIA40:ERV1 oxidizes cysteine residues to cystine disulfide bonds |
| GO:0140318 protein transporter activity | IDA PMID:9822593 Tim9p, an essential partner subunit of Tim10p for the import... | ACCEPT | Summary: IDA annotation for protein transporter activity based on Koehler et al. (1998), who demonstrated that Tim9p is an essential partner of Tim10p for the import of mitochondrial carrier proteins and can be cross-linked to partly translocated carrier proteins. Reason: GO:0140318 directly describes binding a protein and delivering it to a cellular location, and its live ontology comment explicitly names the soluble Tim9-Tim10 shuttle as an example. The complementary GO:0140309 NEW annotation captures the holdase/anti-aggregation dimension rather than replacing this transporter activity. Supporting Evidence: PMID:9822593 Tim9p can be cross-linked to a partly translocated carrier protein. PMID:9822593 Tim9p is a new subunit of the TIM machinery that guides hydrophobic inner membrane proteins across the aqueous intermembrane space. |
| GO:0140318 protein transporter activity | IGI PMID:9822593 Tim9p, an essential partner subunit of Tim10p for the import... | ACCEPT | Summary: IGI annotation for protein transporter activity from the same study. Tim9p function was shown through genetic interaction with Tim10p (Ser67Cys mutation in Tim9p suppresses temperature-sensitive growth defects of tim10-1 and tim12-1 mutants). Reason: The genetic interaction data (suppression of tim10-1 and tim12-1) support the functional involvement of TIM9 in protein transport, consistent with its role in the carrier import pathway. Supporting Evidence: PMID:9822593 A Ser67-->Cys67 mutation in Tim9p suppresses the temperature-sensitive growth defect of tim10-1 and tim12-1 mutants. |
| GO:0005739 mitochondrion | HDA PMID:24769239 Quantitative variations of the mitochondrial proteome and ph... | ACCEPT | Summary: HDA (high-throughput direct assay) annotation for mitochondrial localization based on Renvoise et al. (2014), a quantitative proteomic study of isolated yeast mitochondria. TIM9 was identified by mass spectrometry in mitochondrial fractions. Reason: Proteomic identification of TIM9 in mitochondrial fractions is consistent with all other evidence for its mitochondrial localization. Supporting Evidence: PMID:24769239 Label free quantitative analysis of protein accumulation revealed significant variation of 176 mitochondrial proteins |
| GO:0005739 mitochondrion | HDA PMID:16823961 Toward the complete yeast mitochondrial proteome: multidimen... | ACCEPT | Summary: HDA annotation for mitochondrial localization based on Reinders et al. (2006), a comprehensive mitochondrial proteomics study that identified 851 proteins in yeast mitochondria using multidimensional LC-MS/MS approaches. Reason: Mass spectrometry identification of TIM9 in purified mitochondria confirms its mitochondrial localization. The UniProt entry cites this study for subcellular location. Supporting Evidence: PMID:16823961 A total of 851 different proteins (PROMITO dataset) were identified by use of multidimensional LC-MS/MS, 1D-SDS-PAGE combined with nano-LC-MS/MS and 2D-PAGE with subsequent MALDI-mass fingerprinting. |
| GO:0042719 mitochondrial intermembrane space chaperone complex | IDA PMID:9822593 Tim9p, an essential partner subunit of Tim10p for the import... | ACCEPT | Summary: IDA annotation for membership in the mitochondrial IMS chaperone complex (the soluble Tim9-Tim10 70 kDa complex) based on Koehler et al. (1998), who co-purified Tim9p and Tim10p and showed co-immunoprecipitation. Reason: TIM9 is a canonical and essential subunit of the Tim9-Tim10 IMS chaperone complex. This is a core annotation. Supporting Evidence: PMID:9822593 Most of Tim9p is associated with Tim10p in a soluble 70 kDa complex. Tim9p and Tim10p co-purify in successive chromatographic fractionations and co-immunoprecipitated with each other. |
| GO:0042719 mitochondrial intermembrane space chaperone complex | IDA PMID:9889188 Tim9, a new component of the TIM22.54 translocase in mitocho... | ACCEPT | Summary: IDA annotation for IMS chaperone complex membership based on Adam et al. (1999), who independently identified Tim9 and showed it forms distinct complexes with Tim10 and Tim12. Reason: Independent confirmation of TIM9 as a component of the IMS chaperone complex. Supporting Evidence: PMID:9889188 Tim9 is located in the mitochondrial intermembrane space and is organized into two distinct hetero-oligomeric assemblies with Tim10 and Tim12. One complex contains Tim9 and Tim10. |
| GO:0042721 TIM22 mitochondrial import inner membrane insertion complex | IDA PMID:9495346 Carrier protein import into mitochondria mediated by the int... | ACCEPT | Summary: IDA annotation for TIM22 complex membership based on Sirrenberg et al. (1998), who showed that Tim10 and Tim12 interact with precursors and are part of a complex with Tim22 that mediates inner membrane insertion. Tim9 is annotated to this complex, but the cached paper is abstract-only and foregrounds Tim10 and Tim12 rather than Tim9, so it cannot by itself verify the TIM9 assay; subsequent direct studies place Tim9 in the TIM22-associated 300 kDa complex (PMID:9822593, PMID:9889188). Reason: TIM9 is a peripheral TIM22-pathway component via its association with Tim10 and Tim12. The annotation is independently supported by PMID:9822593, while the abstract-only original reference is not overinterpreted or treated as disproving the curator's inaccessible full-text evidence. Supporting Evidence: PMID:9495346 Tim10 and Tim12 are found in a complex with Tim22, which takes over the precursor and mediates its membrane-potential-dependent insertion into the inner membrane. PMID:9822593 A small fraction of Tim9p is bound to the outer face of the inner membrane in a 300 kDa complex whose other subunits include Tim54p, Tim22p, Tim12p and Tim10p. |
| GO:0045039 protein insertion into mitochondrial inner membrane | IDA PMID:10469659 Different import pathways through the mitochondrial intermem... | ACCEPT | Summary: IDA annotation based on Leuenberger et al. (1999), who showed that the Tim9p-Tim10p complex mediates the import of a specific subset of integral inner membrane proteins and can transfer these proteins to one of three different membrane insertion sites. Reason: This study extends the known role of TIM9 beyond carrier proteins to additional inner membrane protein substrates, demonstrating the breadth of its involvement in protein insertion into the inner membrane. Supporting Evidence: PMID:10469659 the two 70 kDa complexes each mediate the import of a different subset of integral inner membrane proteins and that they can transfer these proteins to one of three different membrane insertion sites: the TIM22 complex, the TIM23 complex or an as yet uncharacterized insertion site. |
| GO:0045039 protein insertion into mitochondrial inner membrane | IDA PMID:9822593 Tim9p, an essential partner subunit of Tim10p for the import... | ACCEPT | Summary: IDA annotation based on Koehler et al. (1998), the original identification of Tim9p as an essential partner of Tim10p for import of mitochondrial carrier proteins. Reason: Foundational study demonstrating TIM9 involvement in carrier protein import. Tim9p was shown to be cross-linkable to partly translocated carrier proteins. Supporting Evidence: PMID:9822593 Tim9p can be cross-linked to a partly translocated carrier protein. |
| GO:0045039 protein insertion into mitochondrial inner membrane | IGI PMID:9822593 Tim9p, an essential partner subunit of Tim10p for the import... | ACCEPT | Summary: IGI annotation based on genetic interaction evidence from Koehler et al. (1998). The Ser67Cys mutation in Tim9p suppresses ts growth defects of tim10-1 and tim12-1, demonstrating genetic interaction within the carrier import pathway. Reason: Genetic suppression data (Tim9p Ser67Cys suppresses tim10-1 and tim12-1) support the functional involvement of TIM9 in the carrier protein import pathway. Supporting Evidence: PMID:9822593 A Ser67-->Cys67 mutation in Tim9p suppresses the temperature-sensitive growth defect of tim10-1 and tim12-1 mutants. |
| GO:0045039 protein insertion into mitochondrial inner membrane | IMP PMID:9889188 Tim9, a new component of the TIM22.54 translocase in mitocho... | ACCEPT | Summary: IMP annotation based on Adam et al. (1999), who demonstrated that Tim9 is an essential protein and that the TIM9.10 complex mediates partial translocation of carrier proteins. Reason: Mutant phenotype evidence (essentiality and translocation defects) supports the role of TIM9 in protein insertion into the mitochondrial inner membrane. Supporting Evidence: PMID:9889188 We have identified Tim9, a new component of the TIM22.54 import machinery, which mediates transport of proteins into the inner membrane of mitochondria. Tim9, an essential protein of Saccharomyces cerevisiae |
| GO:0051082 unfolded protein binding | IDA PMID:12138093 Assembly of Tim9 and Tim10 into a functional chaperone. | MODIFY | Summary: IDA annotation for unfolded protein binding based on Vial et al. (2002), who demonstrated that the reconstituted Tim9-Tim10 complex binds to the physiological substrate ADP/ATP carrier (AAC) and displays chaperone activity in refolding the model substrate firefly luciferase. TIM9 contributes to the canonical carrier-holdase complex: it binds unfolded/hydrophobic protein precursors AND escorts them across the mitochondrial IMS from the TOM complex to the TIM22 complex. GO:0051082 "unfolded protein binding" captures only the binding aspect but misses the essential carrier/escort function. GO:0140309 "unfolded protein holdase activity" was created specifically for this class of proteins (go-ontology#30552) and is the correct replacement term. Reason: TIM9 (as part of the Tim9-Tim10 complex) contributes to a textbook carrier-holdase: it binds unfolded hydrophobic precursor proteins AND escorts them between cellular compartments (principally from TOM to TIM22 across the IMS). GO:0051082 "unfolded protein binding" captures only the binding aspect. GO:0140309 "unfolded protein holdase activity" was created in Nov 2025 specifically for TIM carrier-holdases (go-ontology#30552). The current definition emphasizes binding an unfolded client, escorting it to an acceptor or location, and preventing aggregation. The Tim9-Tim10 complex acts as a chaperone-like protein that protects hydrophobic precursors from aggregation and guides them through the mitochondrial intermembrane space (UniProt). PMID:12138093 shows the reconstituted complex is functional because it binds AAC and displays chaperone activity. Proposed replacements: unfolded protein holdase activity Supporting Evidence: PMID:12138093 the reconstituted TIM10 complex is functional because it bound to the physiological substrate ADP/ATP carrier and displayed chaperone activity in refolding the model substrate firefly luciferase. PMID:9822593 Tim9p is a new subunit of the TIM machinery that guides hydrophobic inner membrane proteins across the aqueous intermembrane space. UniProt:O74700 Acts as a chaperone-like protein that protects the hydrophobic precursors from aggregation and guide them through the mitochondrial intermembrane space. file:yeast/TIM9/TIM9-deep-research-falcon.md Tim9 acts as a **chaperone/escort factor** that prevents aggregation of hydrophobic precursors in the aqueous IMS and transfers them from **TOM** to **TIM22**. file:yeast/TIM9/TIM9-deep-research-falcon.md TIM9·10 recognizes hydrophobic clients primarily through a **hydrophobic cleft** formed by chaperone helices |
| GO:0140309 unfolded protein holdase activity | IDA PMID:12138093 Assembly of Tim9 and Tim10 into a functional chaperone. | NEW | Summary: NEW annotation. TIM9 (as part of the Tim9-Tim10 hexameric complex) is the canonical carrier-holdase for which GO:0140309 was created (go-ontology#30552). The Tim9-Tim10 complex binds unfolded hydrophobic transmembrane protein precursors and escorts them from the TOM complex across the IMS to the TIM22 complex for inner membrane insertion. Small Tim complexes have also been implicated in some SAM-directed beta-barrel precursor delivery, but that is not required for this NEW assertion. Vial et al. (2002, PMID:12138093) showed the reconstituted Tim9-Tim10 complex binds the ADP/ATP carrier substrate and displays chaperone activity. Multiple studies demonstrate the carrier/escort function: the complex guides hydrophobic proteins through the aqueous IMS (PMID:9822593), mediates partial translocation of carrier proteins (PMID:9889188), and can transfer substrates to multiple insertion sites (PMID:10469659). Reason: GO:0140309 "unfolded protein holdase activity" was created specifically for TIM carrier-holdases. TIM9 is the founding member of this functional class. This term is not currently annotated to TIM9 in GOA despite being the most precise MF term for its chaperone function. It should be added as a new annotation. Supporting Evidence: PMID:12138093 the reconstituted TIM10 complex is functional because it bound to the physiological substrate ADP/ATP carrier and displayed chaperone activity in refolding the model substrate firefly luciferase. PMID:9822593 Tim9p is a new subunit of the TIM machinery that guides hydrophobic inner membrane proteins across the aqueous intermembrane space. PMID:9889188 The TIM9.10 complex is more abundant than the TIM9.10.12 complex and mediates partial translocation of mitochondrial carriers proteins across the outer membrane. PMID:10469659 the two 70 kDa complexes each mediate the import of a different subset of integral inner membrane proteins and that they can transfer these proteins to one of three different membrane insertion sites file:yeast/TIM9/TIM9-deep-research-falcon.md Tim9 is best viewed as a **specialized, soluble IMS holdase chaperone** file:yeast/TIM9/TIM9-deep-research-falcon.md Small TIMs can also assist trafficking of some outer-membrane β-barrel proteins toward the SAM complex |
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Download this section (compressed HTML)Q: What proportion of TIM9 client handling is directed to TIM22 versus SAM in vivo, and which individual beta-barrel precursors require the Tim9-Tim10 complex?
Q: How do Tim9's structural contribution to the heterohexamer and Tim10's stronger isolated AAC-binding capacity partition client recognition within the intact complex?
Experiment: Perform time-resolved crosslinking of endogenous Tim9 and Tim10 during synchronized import of carrier and beta-barrel precursor panels, quantifying handoff to TIM22 or SAM.
Hypothesis: Tim9-Tim10 directly handles both client classes, but TIM22-directed carrier delivery is the dominant and most robust TIM9-dependent route.
Experiment: Reconstitute Tim9-Tim10 complexes carrying interface-preserving Tim9 client-contact mutations and measure complex assembly, aggregation suppression, and precursor delivery.
Hypothesis: Tim9 contributes client-contact surfaces beyond its structural role, even though isolated Tim10 shows stronger AAC binding than isolated Tim9.
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