scholarly journals A New Function in Translocation for the Mitochondrial i-AAA Protease Yme1: Import of Polynucleotide Phosphorylase into the Intermembrane Space

2006 ◽  
Vol 26 (22) ◽  
pp. 8488-8497 ◽  
Author(s):  
Robert N. Rainey ◽  
Jenny D. Glavin ◽  
Hsiao-Wen Chen ◽  
Samuel W. French ◽  
Michael A. Teitell ◽  
...  
Keyword(s):  
Protein Translocation ◽  
Rna Degradation ◽  
Mechanistic Study ◽  
Intermembrane Space ◽  
Polynucleotide Phosphorylase ◽  
Cellular Effects ◽  
The Matrix ◽  
Mitochondrial Intermembrane Space ◽  
Processing Peptidase

ABSTRACT Polynucleotide phosphorylase (PNPase) is an exoribonuclease and poly(A) polymerase postulated to function in the cytosol and mitochondrial matrix. Prior overexpression studies resulted in PNPase localization to both the cytosol and mitochondria, concurrent with cytosolic RNA degradation and pleiotropic cellular effects, including growth inhibition and apoptosis, that may not reflect a physiologic role for endogenous PNPase. We therefore conducted a mechanistic study of PNPase biogenesis in the mitochondrion. Interestingly, PNPase is localized to the intermembrane space by a novel import pathway. PNPase has a typical N-terminal targeting sequence that is cleaved by the matrix processing peptidase when PNPase engaged the TIM23 translocon at the inner membrane. The i-AAA protease Yme1 mediated translocation of PNPase into the intermembrane space but did not degrade PNPase. In a yeast strain deleted for Yme1 and expressing PNPase, nonimported PNPase accumulated in the cytosol, confirming an in vivo role for Yme1 in PNPase maturation. PNPase localization to the mitochondrial intermembrane space suggests a unique role distinct from its highly conserved function in RNA processing in chloroplasts and bacteria. Furthermore, Yme1 has a new function in protein translocation, indicating that the intermembrane space harbors diverse pathways for protein translocation.

Computational studies of the mitochondrial carrier family SLC25. Present status and future perspectives

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Andrea Pasquadibisceglie ◽  
Fabio Polticelli
Keyword(s):  
Transport Mechanism ◽  
Experimental Studies ◽  
Intermembrane Space ◽  
Mitochondrial Carrier ◽  
Mitochondrial Homeostasis ◽  
The Matrix ◽  
Mitochondrial Carrier Family ◽  
Mitochondrial Intermembrane Space ◽  
Computational Analyses

Abstract The members of the mitochondrial carrier family, also known as solute carrier family 25 (SLC25), are transmembrane proteins involved in the translocation of a plethora of small molecules between the mitochondrial intermembrane space and the matrix. These transporters are characterized by three homologous domains structure and a transport mechanism that involves the transition between different conformations. Mutations in regions critical for these transporters’ function often cause several diseases, given the crucial role of these proteins in the mitochondrial homeostasis. Experimental studies can be problematic in the case of membrane proteins, in particular concerning the characterization of the structure–function relationships. For this reason, computational methods are often applied in order to develop new hypotheses or to support/explain experimental evidence. Here the computational analyses carried out on the SLC25 members are reviewed, describing the main techniques used and the outcome in terms of improved knowledge of the transport mechanism. Potential future applications on this protein family of more recent and advanced in silico methods are also suggested.

scholarly journals Mia40 is a trans-site receptor that drives protein import into the mitochondrial intermembrane space by hydrophobic substrate binding

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Valentina Peleh ◽  
Emmanuelle Cordat ◽  
Johannes M Herrmann
Keyword(s):  
Disulfide Bonds ◽  
Protein Import ◽  
Protein Translocation ◽  
Hydrophobic Interactions ◽  
Substrate Binding ◽  
Intermembrane Space ◽  
Cysteine Motif ◽  
Mitochondrial Intermembrane Space ◽  
Necessary And Sufficient ◽  
Yeast Mutants

Many proteins of the mitochondrial IMS contain conserved cysteines that are oxidized to disulfide bonds during their import. The conserved IMS protein Mia40 is essential for the oxidation and import of these proteins. Mia40 consists of two functional elements: an N-terminal cysteine-proline-cysteine motif conferring substrate oxidation, and a C-terminal hydrophobic pocket for substrate binding. In this study, we generated yeast mutants to dissect both Mia40 activities genetically and biochemically. Thereby we show that the substrate-binding domain of Mia40 is both necessary and sufficient to promote protein import, indicating that trapping by Mia40 drives protein translocation. An oxidase-deficient Mia40 mutant is inviable, but can be partially rescued by the addition of the chemical oxidant diamide. Our results indicate that Mia40 predominantly serves as a trans-site receptor of mitochondria that binds incoming proteins via hydrophobic interactions thereby mediating protein translocation across the outer membrane by a ‘holding trap’ rather than a ‘folding trap’ mechanism.

scholarly journals Mutational Analysis of the Saccharomyces cerevisiae Cytochrome c Oxidase Assembly Protein Cox11p

2006 ◽  
Vol 5 (3) ◽  
pp. 568-578 ◽  
Author(s):  
Graham S. Banting ◽  
D. Moira Glerum
Keyword(s):  
Hydrogen Peroxide ◽  
Cytochrome Oxidase ◽  
Solution Structure ◽  
Sinorhizobium Meliloti ◽  
Mutational Analysis ◽  
Intermembrane Space ◽  
Copper Binding ◽  
Inner Mitochondrial Membrane ◽  
The Matrix

ABSTRACT Cox11p is an integral protein of the inner mitochondrial membrane that is essential for cytochrome c oxidase assembly. The bulk of the protein is located in the intermembrane space and displays high levels of evolutionary conservation. We have analyzed a collection of site-directed and random cox11 mutants in an effort to further define essential portions of the molecule. Of the alleles studied, more than half had no apparent effect on Cox11p function. Among the respiration deficiency-encoding alleles, we identified three distinct phenotypes, which included a set of mutants with a misassembled or partially assembled cytochrome oxidase, as indicated by a blue-shifted cytochrome aa 3 peak. In addition to the shifted spectral signal, these mutants also display a specific reduction in the levels of subunit 1 (Cox1p). Two of these mutations are likely to occlude a surface pocket behind the copper-binding domain in Cox11p, based on analogy with the Sinorhizobium meliloti Cox11 solution structure, thereby suggesting that this pocket is crucial for Cox11p function. Sequential deletions of the matrix portion of Cox11p suggest that this domain is not functional beyond the residues involved in mitochondrial targeting and membrane insertion. In addition, our studies indicate that Δcox11, like Δsco1, displays a specific hypersensitivity to hydrogen peroxide. Our studies provide the first evidence at the level of the cytochrome oxidase holoenzyme that Cox1p is the in vivo target for Cox11p and suggest that Cox11p may also have a role in the response to hydrogen peroxide exposure.

scholarly journals Tim23–Tim50 pair coordinates functions of translocators and motor proteins in mitochondrial protein import

2009 ◽  
Vol 184 (1) ◽  
pp. 129-141 ◽  
Author(s):  
Yasushi Tamura ◽  
Yoshihiro Harada ◽  
Takuya Shiota ◽  
Koji Yamano ◽  
Kazuaki Watanabe ◽  
...  
Keyword(s):  
Motor Proteins ◽  
Protein Import ◽  
Protein Translocation ◽  
Mitochondrial Protein ◽  
Intermembrane Space ◽  
Mitochondrial Protein Import ◽  
Inner Membrane ◽  
The Matrix ◽  
General Protein ◽  
Mitochondrial Hsp70

Mitochondrial protein traffic requires coordinated operation of protein translocator complexes in the mitochondrial membrane. The TIM23 complex translocates and inserts proteins into the mitochondrial inner membrane. Here we analyze the intermembrane space (IMS) domains of Tim23 and Tim50, which are essential subunits of the TIM23 complex, in these functions. We find that interactions of Tim23 and Tim50 in the IMS facilitate transfer of precursor proteins from the TOM40 complex, a general protein translocator in the outer membrane, to the TIM23 complex. Tim23–Tim50 interactions also facilitate a late step of protein translocation across the inner membrane by promoting motor functions of mitochondrial Hsp70 in the matrix. Therefore, the Tim23–Tim50 pair coordinates the actions of the TOM40 and TIM23 complexes together with motor proteins for mitochondrial protein import.

scholarly journals In Vivo Structure-Function Analysis and Redox Interactomes of Leishmania tarentolae Erv

2021 ◽  
Author(s):  
Gino L. Turra ◽  
Linda Liedgens ◽  
Frederik Sommer ◽  
Luzia Schneider ◽  
David Zimmer ◽  
...  
Keyword(s):  
Protein Import ◽  
Function Analysis ◽  
Mitochondrial Protein ◽  
Intermembrane Space ◽  
Mitochondrial Protein Import ◽  
Redox Biology ◽  
Oxidative Protein Folding ◽  
Mitochondrial Intermembrane Space ◽  
New Research

The discovery of the redox proteins Mia40/CHCHD4 and Erv1/ALR, as well as the elucidation of their relevance for oxidative protein folding in the mitochondrial intermembrane space of yeast and mammals, founded a new research topic in redox biology and mitochondrial protein import. The lack of Mia40/CHCHD4 in protist lineages raises fundamental and controversial questions regarding the conservation and evolution of this essential pathway.

scholarly journals A new assembly pathway in the mitochondrial intermembrane space for mammalian polynucleotide phosphorylase

2006 ◽  
Vol 20 (5) ◽  
Author(s):  
Jenny D Glavin ◽  
Robert N. Rainey ◽  
Hsiao‐Wen Chen ◽  
Michael A. Teitell ◽  
Carla M. Koehler
Keyword(s):  
Intermembrane Space ◽  
Polynucleotide Phosphorylase ◽  
Assembly Pathway ◽  
Mitochondrial Intermembrane Space

scholarly journals The mitochondrial intermembrane space: the most constricted mitochondrial sub-compartment with the largest variety of protein import pathways

Open Biology ◽  
2021 ◽  
Vol 11 (3) ◽  
Author(s):  
Ruairidh Edwards ◽  
Ross Eaglesfield ◽  
Kostas Tokatlidis
Keyword(s):  
Protein Import ◽  
Atp Hydrolysis ◽  
Current Knowledge ◽  
Intermembrane Space ◽  
Inner Membrane ◽  
Mitochondrial Proteome ◽  
Normal Site ◽  
The Matrix ◽  
Human Disorders ◽  
Mitochondrial Intermembrane Space

The mitochondrial intermembrane space (IMS) is the most constricted sub-mitochondrial compartment, housing only about 5% of the mitochondrial proteome, and yet is endowed with the largest variability of protein import mechanisms. In this review, we summarize our current knowledge of the major IMS import pathway based on the oxidative protein folding pathway and discuss the stunning variability of other IMS protein import pathways. As IMS-localized proteins only have to cross the outer mitochondrial membrane, they do not require energy sources like ATP hydrolysis in the mitochondrial matrix or the inner membrane electrochemical potential which are critical for import into the matrix or insertion into the inner membrane. We also explore several atypical IMS import pathways that are still not very well understood and are guided by poorly defined or completely unknown targeting peptides. Importantly, many of the IMS proteins are linked to several human diseases, and it is therefore crucial to understand how they reach their normal site of function in the IMS. In the final part of this review, we discuss current understanding of how such IMS protein underpin a large spectrum of human disorders.

Mutations in a 19-amino-acid hydrophobic region of the yeast cytochrome c1 presequence prevent sorting to the mitochondrial intermembrane space

1992 ◽  
Vol 12 (10) ◽  
pp. 4677-4686
Author(s):  
R E Jensen ◽  
S Schmidt ◽  
R J Mark
Keyword(s):  
Amino Acid ◽  
Intermediate Form ◽  
Intermembrane Space ◽  
Protein Amino Acid ◽  
Hydrophobic Region ◽  
Yeast Saccharomyces Cerevisiae ◽  
Cytochrome C1 ◽  
The Matrix ◽  
Hydrophobic Amino Acids ◽  
Mitochondrial Intermembrane Space

Most mitochondrial proteins destined for the intermembrane space (IMS) carry in their presequence information for localization to the IMS in addition to information for their import. By selecting for mutants in the yeast Saccharomyces cerevisiae that mislocalize an IMS-targeted fusion protein, we identified mutations in the IMS sorting signal of the cytochrome c1 protein. Amino acid substitutions or deletions in a stretch of 19 hydrophobic amino acids of the cytochrome c1 presequence resulted in accumulation of the intermediate form of the cytochrome c1 protein in the matrix. In some cases, the accumulated intermediate appeared to be slowly exported from the matrix, across the inner membrane to the IMS. Our results support the hypothesis that the cytochrome c1 precursor is normally imported completely into the matrix and then exported to the IMS.

scholarly journals The mitochondrial carrier pathway transports non-canonical substrates with an odd number of transmembrane segments

BMC Biology ◽  
2020 ◽  
Vol 18 (1) ◽  
Author(s):  
Heike Rampelt ◽  
Iva Sucec ◽  
Beate Bersch ◽  
Patrick Horten ◽  
Inge Perschil ◽  
...  
Keyword(s):  
Intermembrane Space ◽  
Inner Mitochondrial Membrane ◽  
Inner Membrane ◽  
Mitochondrial Carrier ◽  
Transmembrane Segments ◽  
Mitochondrial Inner Membrane ◽  
N Terminus ◽  
The Matrix ◽  
Mitochondrial Carrier Family ◽  
Mitochondrial Intermembrane Space

Abstract Background The mitochondrial pyruvate carrier (MPC) plays a central role in energy metabolism by transporting pyruvate across the inner mitochondrial membrane. Its heterodimeric composition and homology to SWEET and semiSWEET transporters set the MPC apart from the canonical mitochondrial carrier family (named MCF or SLC25). The import of the canonical carriers is mediated by the carrier translocase of the inner membrane (TIM22) pathway and is dependent on their structure, which features an even number of transmembrane segments and both termini in the intermembrane space. The import pathway of MPC proteins has not been elucidated. The odd number of transmembrane segments and positioning of the N-terminus in the matrix argues against an import via the TIM22 carrier pathway but favors an import via the flexible presequence pathway. Results Here, we systematically analyzed the import pathways of Mpc2 and Mpc3 and report that, contrary to an expected import via the flexible presequence pathway, yeast MPC proteins with an odd number of transmembrane segments and matrix-exposed N-terminus are imported by the carrier pathway, using the receptor Tom70, small TIM chaperones, and the TIM22 complex. The TIM9·10 complex chaperones MPC proteins through the mitochondrial intermembrane space using conserved hydrophobic motifs that are also required for the interaction with canonical carrier proteins. Conclusions The carrier pathway can import paired and non-paired transmembrane helices and translocate N-termini to either side of the mitochondrial inner membrane, revealing an unexpected versatility of the mitochondrial import pathway for non-cleavable inner membrane proteins.

scholarly journals Residues of Tim44 Involved in both Association with the Translocon of the Inner Mitochondrial Membrane and Regulation of Mitochondrial Hsp70 Tethering

2008 ◽  
Vol 28 (13) ◽  
pp. 4424-4433 ◽  
Author(s):  
Dirk Schiller ◽  
Yu Chin Cheng ◽  
Qinglian Liu ◽  
William Walter ◽  
Elizabeth A. Craig
Keyword(s):  
Amino Acid ◽  
Protein Import ◽  
Protein Translocation ◽  
Mitochondrial Protein ◽  
Inner Mitochondrial Membrane ◽  
The Matrix ◽  
Mitochondrial Hsp70 ◽  
Amino Acid Segment

ABSTRACT Translocation of proteins from the cytosol across the mitochondrial inner membrane is driven by the action of the import motor, which is associated with the translocon on the matrix side of the membrane. It is well established that an essential peripheral membrane protein, Tim44, tethers mitochondrial Hsp70 (mtHsp70), the core of the import motor, to the translocon. This Tim44-mtHsp70 interaction, which can be recapitulated in vitro, is destabilized by binding of mtHsp70 to a substrate polypeptide. Here we report that the N-terminal 167-amino-acid segment of mature Tim44 is sufficient for both interaction with mtHsp70 and destabilization of a Tim44-mtHsp70 complex caused by client protein binding. Amino acid alterations within a 30-amino-acid segment affected both the release of mtHsp70 upon peptide binding and the interaction of Tim44 with the translocon. Our results support the idea that Tim44 plays multiple roles in mitochondrial protein import by recruiting Ssc1 and its J protein cochaperone to the translocon and coordinating their interactions to promote efficient protein translocation in vivo.

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