scholarly journals Mitochondrial protein import: biochemical and genetic evidence for interaction of matrix hsp70 and the inner membrane protein MIM44.

1994 ◽  
Vol 127 (6) ◽  
pp. 1547-1556 ◽  
Author(s):  
J Rassow ◽  
A C Maarse ◽  
E Krainer ◽  
M Kübrich ◽  
H Müller ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Binding Proteins ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Growth Defect ◽  
Inner Membrane ◽  
The Matrix ◽  
Inner Membrane Protein ◽  
Polypeptide Chains ◽  
Matrix Heat

The import of preproteins into mitochondria involves translocation of the polypeptide chains through putative channels in the outer and inner membranes. Preprotein-binding proteins are needed to drive the unidirectional translocation of the precursor polypeptides. Two of these preprotein-binding proteins are the peripheral inner membrane protein MIM44 and the matrix heat shock protein hsp70. We report here that MIM44 is mainly exposed on the matrix side, and a fraction of mt-hsp70 is reversibly bound to the inner membrane. Mt-hsp70 binds to MIM44 in a 1:1 ratio, suggesting that mt-hsp70 is localizing to the membrane via its interaction with MIM44. Formation of the complex requires a functional ATPase domain of mt-hsp70. Addition of Mg-ATP leads to dissociation of the complex. Overexpression of mt-hsp70 rescues the protein import defect of mutants in MIM44; conversely, overexpression of MIM44 rescues protein import defects of mt-hsp70 mutants. In addition, yeast strains with conditional mutations in both MIM44 and mt-hsp70 are barely viable, showing a synthetic growth defect compared to strains carrying single mutations. We propose that MIM44 and mt-hsp70 cooperate in translocation of preproteins. By binding to MIM44, mt-hsp70 is recruited at the protein import sites of the inner membrane, and preproteins arriving at MIM44 may be directly handed over to mt-hsp70.

scholarly journals SMS1, a high-copy suppressor of the yeast mas6 mutant, encodes an essential inner membrane protein required for mitochondrial protein import.

1994 ◽  
Vol 5 (5) ◽  
pp. 529-538 ◽  
Author(s):  
K R Ryan ◽  
M M Menold ◽  
S Garrett ◽  
R E Jensen
Keyword(s):  
Membrane Protein ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Mitochondrial Protein Import ◽  
Yeast Saccharomyces Cerevisiae ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Inner Membrane Protein ◽  
Membrane Spanning ◽  
Membrane Spanning Domains

MAS6 encodes an essential inner membrane protein required for mitochondrial protein import in the yeast Saccharomyces cerevisiae (Emtage and Jensen, 1993). To identify new inner membrane import components, we isolated a high-copy suppressor (SMS1) of the mas6-1 mutant. SMS1 encodes a 16.5-kDa protein that contains several potential membrane-spanning domains. The Sms1 protein is homologous to the carboxyl-terminal domain of the Mas6 protein. Like Mas6p, Sms1p is located in the mitochondrial inner membrane and is an essential protein. Depletion of Sms1p from cells causes defects in the import of several mitochondrial precursor proteins, suggesting that Sms1p is a new inner membrane import component. Our observations raise the possibility that Sms1p and Mas6p act together to translocate proteins across the inner membrane.

scholarly journals Mitochondrial Protein Import Motor: Differential Role of Tim44 in the Recruitment of Pam17 and J-Complex to the Presequence Translocase

2008 ◽  
Vol 19 (6) ◽  
pp. 2642-2649 ◽  
Author(s):  
Dana P. Hutu ◽  
Bernard Guiard ◽  
Agnieszka Chacinska ◽  
Dorothea Becker ◽  
Nikolaus Pfanner ◽  
...  
Keyword(s):  
Protein Import ◽  
Mitochondrial Protein ◽  
Adaptor Protein ◽  
Inner Membrane ◽  
Amino Terminal ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Yeast Mutants ◽  
Matrix Heat

The presequence translocase of the mitochondrial inner membrane (TIM23 complex) mediates the import of preproteins with amino-terminal presequences. To drive matrix translocation the TIM23 complex recruits the presequence translocase-associated motor (PAM) with the matrix heat shock protein 70 (mtHsp70) as central subunit. Activity and localization of mtHsp70 are regulated by four membrane-associated cochaperones: the adaptor protein Tim44, the stimulatory J-complex Pam18/Pam16, and Pam17. It has been proposed that Tim44 serves as molecular platform to localize mtHsp70 and the J-complex at the TIM23 complex, but it is unknown how Pam17 interacts with the translocase. We generated conditional tim44 yeast mutants and selected a mutant allele, which differentially affects the association of PAM modules with TIM23. In tim44-804 mitochondria, the interaction of the J-complex with the TIM23 complex is impaired, whereas unexpectedly the binding of Pam17 is increased. Pam17 interacts with the channel protein Tim23, revealing a new interaction site between TIM23 and PAM. Thus, the motor PAM is composed of functional modules that bind to different sites of the translocase. We suggest that Tim44 is not simply a scaffold for binding of motor subunits but plays a differential role in the recruitment of PAM modules to the inner membrane translocase.

scholarly journals Characterization of Mmp37p, aSaccharomyces cerevisiaeMitochondrial Matrix Protein with a Role in Mitochondrial Protein Import

2006 ◽  
Vol 17 (9) ◽  
pp. 4051-4062 ◽  
Author(s):  
Michelle R. Gallas ◽  
Mary K. Dienhart ◽  
Rosemary A. Stuart ◽  
Roy M. Long
Keyword(s):  
Matrix Protein ◽  
Protein Import ◽  
Mitochondrial Protein ◽  
Temperature Sensitive ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Close Proximity ◽  
The Matrix ◽  
Targeting Signal

Many mitochondrial proteins are encoded by nuclear genes and after translation in the cytoplasm are imported via translocases in the outer and inner membranes, the TOM and TIM complexes, respectively. Here, we report the characterization of the mitochondrial protein, Mmp37p (YGR046w) and demonstrate its involvement in the process of protein import into mitochondria. Haploid cells deleted of MMP37 are viable but display a temperature-sensitive growth phenotype and are inviable in the absence of mitochondrial DNA. Mmp37p is located in the mitochondrial matrix where it is peripherally associated with the inner membrane. We show that Mmp37p has a role in the translocation of proteins across the mitochondrial inner membrane via the TIM23-PAM complex and further demonstrate that substrates containing a tightly folded domain in close proximity to their mitochondrial targeting sequences display a particular dependency on Mmp37p for mitochondrial import. Prior unfolding of the preprotein, or extension of the region between the targeting signal and the tightly folded domain, relieves their dependency for Mmp37p. Furthermore, evidence is presented to show that Mmp37 may affect the assembly state of the TIM23 complex. On the basis of these findings, we hypothesize that the presence of Mmp37p enhances the early stages of the TIM23 matrix import pathway to ensure engagement of incoming preproteins with the mtHsp70p/PAM complex, a step that is necessary to drive the unfolding and complete translocation of the preprotein into the matrix.

scholarly journals Evolution of mitochondrial protein import – lessons from trypanosomes

2020 ◽  
Vol 401 (6-7) ◽  
pp. 663-676 ◽  
Author(s):  
André Schneider
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Protein Import ◽  
Outer Membrane Proteins ◽  
Mitochondrial Protein ◽  
Intermembrane Space ◽  
Mitochondrial Protein Import ◽  
System A ◽  
Import Receptors ◽  
Inner Membrane Protein

AbstractThe evolution of mitochondrial protein import and the systems that mediate it marks the boundary between the endosymbiotic ancestor of mitochondria and a true organelle that is under the control of the nucleus. Protein import has been studied in great detail in Saccharomyces cerevisiae. More recently, it has also been extensively investigated in the parasitic protozoan Trypanosoma brucei, making it arguably the second best studied system. A comparative analysis of the protein import complexes of yeast and trypanosomes is provided. Together with data from other systems, this allows to reconstruct the ancestral features of import complexes that were present in the last eukaryotic common ancestor (LECA) and to identify which subunits were added later in evolution. How these data can be translated into plausible scenarios is discussed, providing insights into the evolution of (i) outer membrane protein import receptors, (ii) proteins involved in biogenesis of α-helically anchored outer membrane proteins, and (iii) of the intermembrane space import and assembly system. Finally, it is shown that the unusual presequence-associated import motor of trypanosomes suggests a scenario of how the two ancestral inner membrane protein translocases present in LECA evolved into the single bifunctional one found in extant trypanosomes.

scholarly journals Identification of the essential yeast protein MIM17, an integral mitochondrial inner membrane protein involved in protein import

FEBS Letters ◽  
1994 ◽  
Vol 349 (2) ◽  
pp. 215-221 ◽  
Author(s):  
Ammy C. Maarse ◽  
Jolanda Blom ◽  
Petra Keil ◽  
Nikolaus Pfanner ◽  
Michiel Meijer
Keyword(s):  
Membrane Protein ◽  
Protein Import ◽  
Yeast Protein ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Inner Membrane Protein

scholarly journals A J-protein is an essential subunit of the presequence translocase–associated protein import motor of mitochondria

2003 ◽  
Vol 163 (4) ◽  
pp. 707-713 ◽  
Author(s):  
Kaye N. Truscott ◽  
Wolfgang Voos ◽  
Ann E. Frazier ◽  
Maria Lind ◽  
Yanfeng Li ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Protein Import ◽  
Protein Translocation ◽  
Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
The Matrix ◽  
Inner Membrane Protein ◽  
Mitochondrial Hsp70 ◽  
Essential Subunit ◽  
J Protein

Transport of preproteins into the mitochondrial matrix is mediated by the presequence translocase–associated motor (PAM). Three essential subunits of the motor are known: mitochondrial Hsp70 (mtHsp70); the peripheral membrane protein Tim44; and the nucleotide exchange factor Mge1. We have identified the fourth essential subunit of the PAM, an essential inner membrane protein of 18 kD with a J-domain that stimulates the ATPase activity of mtHsp70. The novel J-protein (encoded by PAM18/YLR008c/TIM14) is required for the interaction of mtHsp70 with Tim44 and protein translocation into the matrix. We conclude that the reaction cycle of the PAM of mitochondria involves an essential J-protein.

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 ROMO1 is a constituent of the human presequence translocase required for YME1L protease import

2018 ◽  
Vol 218 (2) ◽  
pp. 598-614 ◽  
Author(s):  
Frank Richter ◽  
Sven Dennerlein ◽  
Miroslav Nikolov ◽  
Daniel C. Jans ◽  
Nataliia Naumenko ◽  
...  
Keyword(s):  
Membrane Structure ◽  
Protein Import ◽  
Protein Quality ◽  
Functional Characterization ◽  
Mitochondrial Protein ◽  
Mitochondrial Protein Import ◽  
Yeast Saccharomyces Cerevisiae ◽  
Inner Membrane ◽  
Inner Membrane Protein ◽  
General Protein

The mitochondrial presequence translocation machinery (TIM23 complex) is conserved between the yeast Saccharomyces cerevisiae and humans; however, functional characterization has been mainly performed in yeast. Here, we define the constituents of the human TIM23 complex using mass spectrometry and identified ROMO1 as a new translocase constituent with an exceptionally short half-life. Analyses of a ROMO1 knockout cell line revealed aberrant inner membrane structure and altered processing of the GTPase OPA1. We show that in the absence of ROMO1, mitochondria lose the inner membrane YME1L protease, which participates in OPA1 processing and ROMO1 turnover. While ROMO1 is dispensable for general protein import along the presequence pathway, we show that it participates in the dynamics of TIM21 during respiratory chain biogenesis and is specifically required for import of YME1L. This selective import defect can be linked to charge distribution in the unusually long targeting sequence of YME1L. Our analyses establish an unexpected link between mitochondrial protein import and inner membrane protein quality control.

scholarly journals Pam17 Is Required for Architecture and Translocation Activity of the Mitochondrial Protein Import Motor

2005 ◽  
Vol 25 (17) ◽  
pp. 7449-7458 ◽  
Author(s):  
Martin van der Laan ◽  
Agnieszka Chacinska ◽  
Maria Lind ◽  
Inge Perschil ◽  
Albert Sickmann ◽  
...  
Keyword(s):  
Protein Import ◽  
Mitochondrial Protein ◽  
Matrix Proteins ◽  
Stable Complex ◽  
Nucleotide Exchange ◽  
Inner Membrane ◽  
Reading Frame ◽  
Nucleotide Exchange Factor ◽  
The Matrix ◽  
Preprotein Translocation

ABSTRACT Import of mitochondrial matrix proteins involves the general translocase of the outer membrane and the presequence translocase of the inner membrane. The presequence translocase-associated motor (PAM) drives the completion of preprotein translocation into the matrix. Five subunits of PAM are known: the preprotein-binding matrix heat shock protein 70 (mtHsp70), the nucleotide exchange factor Mge1, Tim44 that directs mtHsp70 to the inner membrane, and the membrane-bound complex of Pam16-Pam18 that regulates the ATPase activity of mtHsp70. We have identified a sixth motor subunit. Pam17 (encoded by the open reading frame YKR065c) is anchored in the inner membrane and exposed to the matrix. Mitochondria lacking Pam17 are selectively impaired in the import of matrix proteins and the generation of an import-driving activity of PAM. Pam17 is required for formation of a stable complex between the cochaperones Pam16 and Pam18 and promotes the association of Pam16-Pam18 with the presequence translocase. Our findings suggest that Pam17 is required for the correct organization of the Pam16-Pam18 complex and thus contributes to regulation of mtHsp70 activity at the inner membrane translocation site.

scholarly journals Intramitochondrial sorting of the precursor to yeast cytochrome c oxidase subunit Va.

1993 ◽  
Vol 121 (5) ◽  
pp. 1021-1029 ◽  
Author(s):  
B R Miller ◽  
M G Cumsky
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Mitochondrial Protein ◽  
Inner Membrane ◽  
Oxidase Subunit ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Inner Membrane Protein ◽  
Membrane Spanning ◽  
Mitochondrial Heat Shock Protein

We have continued our studies on the import pathway of the precursor to yeast cytochrome c oxidase subunit Va (pVa), a mitochondrial inner membrane protein. Previous work on this precursor demonstrated that import of pVa is unusually efficient, and that inner membrane localization is directed by a membrane-spanning domain in the COOH-terminal third of the protein. Here we report the results of studies aimed at analyzing the intramitochondrial sorting of pVa, as well as the role played by ancillary factors in import and localization of the precursor. We found that pVa was efficiently imported and correctly sorted in mitochondria prepared from yeast strains defective in the function of either mitochondrial heat shock protein (hsp)60 or hsp70. Under identical conditions the import and sorting of another mitochondrial protein, the precursor to the beta subunit of the F1 ATPase, was completely defective. Consistent with previous results demonstrating that the subunit Va precursor is loosely folded, we found that pVa could be efficiently imported into mitochondria after translation in wheat germ extracts. This results suggests that normal levels of extramitochondrial hsp70 are also not required for import of the protein. The results of this study enhance our understanding of the mechanism by which pVa is routed to the mitochondrial inner membrane. They suggest that while the NH2 terminus of pVa is exposed to the matrix and processed by the matrix metalloprotease, the protein remains anchored to the inner membrane before being assembled into a functional holoenzyme complex.

Sign in / Sign up

Export Citation Format

Share Document