Origin of a chloroplast protein importer

Bölter, Bettina; Soll, Jürgen; Schulz, Alexander; Hinnah, Silke C.; Wagner, Richard

doi:10.1073/pnas.95.26.15831

Cited by 144 publications

(101 citation statements)

References 45 publications

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“…Interestingly, Hmp35 is also predicted to be a predominantly ␤-structured protein with a 47% ␤-sheet content, which is comparable with the 45% predicted and the 48 -52% determined experimentally for MspA (53). The Toc75 protein translocase found in chloroplasts likewise exists as a ␤-barrel (54). Furthermore, the ␤-barrel structure is found in all outer membrane proteins in Gramnegative bacteria and a large number of outer membrane proteins in organelles of symbiotic origin, mitochondria, and chloroplasts.…”

Section: Discussionsupporting

confidence: 59%

Trichomonas vaginalis Hmp35, a Putative Pore-forming Hydrogenosomal Membrane Protein, Can Form a Complex in Yeast Mitochondria

Dyall¹,

Lester

Schneider

et al. 2003

Journal of Biological Chemistry

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An abundant integral membrane protein, Hmp35, has been isolated from hydrogenosomes of Trichomonas vaginalis. This protein has no known homologue and exists as a stable 300-kDa complex, termed HMP35, in membranes of the hydrogenosome. By using blue native gel electrophoresis, we found the HMP35 complex to be stable in 2 M NaCl and up to 5 M urea. The endogenous Hmp35 protein was largely protease-resistant. The protein has a predominantly ␤-sheet structure and predicted transmembrane domains that may form a pore. Interestingly, the protein has a high number of cysteine residues, some of which are arranged in motifs that resemble the RING finger, suggesting that they could be coordinating zinc or another divalent cation. Our data show that Hmp35 forms one intramolecular but no intermolecular disulfide bonds. We have isolated the HMP35 complex by expressing a His-tagged Hmp35 protein in vivo followed by purification with nickel-agarose beads. The purified 300-kDa complex consists of mostly Hmp35 with lesser amounts of 12-, 25-27-, and 32-kDa proteins. The stoichiometry of proteins in the complex indicates that Hmp35 exists as an oligomer. Hmp35 can be targeted heterologously into yeast mitochondria, despite the lack of homology with any yeast protein, demonstrating the compatibility of mitochondrial and hydrogenosomal protein translocation machineries.Trichomonas vaginalis is a deep-branching protist that lacks archetypal eukaryotic "aerobic" organelles, specifically mitochondria and peroxisomes. This microaerophilic human-infective parasite carries out fermentative carbohydrate metabolism within hydrogenosomes. Hydrogenosomes are bounded by double membranes and produce ATP by substrate level phosphorylation (1). Hydrogenosomes are also found in certain chytrids, ciliates, and fungi, in lineages that are phylogenetically distant to the Parabasalian lineage to which trichomonads belong (1-4). Currently, several lines of evidence support a common endosymbiotic ancestry for hydrogenosomes and mitochondria, despite their distinct metabolic pathways (3,5,6). Although the origin of hydrogenosomes within ciliate and fungi lineages is debated, these lineages branch with mitochondria-containing groups and the hydrogenosomes confined therein exhibit strong similarity to ciliate and fungal mitochondria (7,8).Trichomonad hydrogenosomes, on the other hand, are markedly less similar to mitochondria. These organelles lack a genome (9) which would have provided a means to investigate their endosymbiotic origin as has been elegantly and convincingly done for mitochondria (10). In lieu of this, we and others have attempted to define the relationship between trichomonad hydrogenosomes and mitochondria by examining the origin of their chaperonins, metabolic enzymes, and membrane proteins. Chaperonin genes, specifically heat shock protein (Hsp) 1 70, cpn60,, and the IscS enzyme, involved in FeS cluster formation (15) appear to have a mitochondrial origin. However, analyses of metabolic enzymes such as hydrogenase (16,17), which is typi...

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Section: Discussionsupporting

confidence: 59%

Trichomonas vaginalis Hmp35, a Putative Pore-forming Hydrogenosomal Membrane Protein, Can Form a Complex in Yeast Mitochondria

Dyall¹,

Lester

Schneider

et al. 2003

Journal of Biological Chemistry

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“…However, no convincing evidence for multimer formation has been obtained for either ShlB or FhaC, thus contrasting with eukaryotic Omp85-like proteins (15, 16). These findings have led others to suggest that the tetrameric Omp85 protein translocation system in eukaryotes may have evolved from more primitive, monomeric bacterial Omp85 homologs (3,5).In this study, we examined Haemophilus influenzae HMW1B, an OM protein that has been demonstrated to be critical for the secretion of the H. influenzae HMW1 adhesin across the OM and is considered to be a member of the TPS pathway, like ShlB and FhaC (12,17). We found that HMW1B exhibits heat-modifiable mobility, a hallmark of ␤-barrel proteins.…”

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Evidence for conservation of architecture and physical properties of Omp85-like proteins throughout evolution

Surana

Grass

Hardy

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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Omp85-like proteins represent a family of proteins involved in protein translocation, and they are present in all domains of life, except archaea. In eukaryotes, Omp85-like proteins have been demonstrated to form tetrameric pore-forming complexes that interact directly with their substrate proteins. Studies performed with bacterial Omp85-like proteins have demonstrated pore-forming activity but no evidence of multimerization. In this article, we characterize the Haemophilus influenzae HMW1B protein, an Omp85-like protein that has been demonstrated to be critical for secretion of the H. influenzae HMW1 adhesin. Analysis of purified protein by biochemical and electron microscopic techniques revealed that HMW1B forms a tetramer. Examination using liposomeswelling assays demonstrated that HMW1B has pore-forming activity, with a pore size of Ϸ2.7 nm. Far-Western blot analysis established that HMW1B interacts with the N terminus of HMW1. These results provide evidence that a bacterial Omp85-like protein forms a tetramer and interacts directly with a substrate protein, suggesting that the architecture and physical properties of Omp85-like proteins have been conserved throughout evolution. O mp85-like proteins represent a large family defined by phylogenetic relatedness, secondary-structure predictions, and a role in protein translocation (1-5). Members of this family are present in diverse organisms across the evolutionary spectrum, including bacteria, fungi, plants, and animals (1, 4). The widespread nature of Omp85-like proteins underscores their critical role in cellular processes, highlighted by the fact that many are required for cell viability (3,4,(6)(7)(8)(9). Although all Omp85-like proteins share sequence homology, they can be grouped into two classes based on the specific role that they play in protein translocation. One class is typified by Saccharomyces cerevisiae Sam50 and Neurospora crassa Tob55, which are chiefly involved in insertion of ␤-barrel proteins into the outer membrane (OM) of mitochondria (6, 9). The second class is epitomized by the chloroplast protein Toc75, which is involved in secretion of proteins across a membrane (10, 11). Interestingly, both classes of Omp85-like proteins are present in Gramnegative bacteria. Neisseria meningitidis Omp85, the namesake of this family of proteins, has been demonstrated recently to be critical for the insertion of proteins into the bacterial OM (7). Other Gram-negative bacterial Omp85-like proteins, including Serratia marcescens ShlB and Bordetella pertussis FhaC [and other members of the so-called two-partner secretion (TPS) pathway], have been demonstrated to function in protein export across the OM (12).The high level of amino acid homology among Omp85-like proteins from bacteria to humans raises the possibility that members of this family have similar structures. Although no crystal structures exist so far for Omp85-like proteins, several of these proteins have been subjected to biochemical studies of overall architecture. In eukaryotes, both classes...

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“…Because homologs exist in extant cyanobacteria (one of which was shown to be essential for viability), these proteins are postulated to be derived from a common ancestor in the original endosymbiont (Bö lter et al, 1998;Reumann et al, 1999Reumann et al, , 2005. Detailed phylogenetic analyses suggested that Toc75 and OEP80 diverged early in the evolution of chloroplasts (Inoue and Potter, 2004).…”

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The Omp85-Related Chloroplast Outer Envelope Protein OEP80 Is Essential for Viability in Arabidopsis

et al. 2008

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b-Barrel proteins of the Omp85 (Outer membrane protein, 85 kD) superfamily exist in the outer membranes of Gram-negative bacteria, mitochondria, and chloroplasts. Prominent Omp85 proteins in bacteria and mitochondria mediate biogenesis of other b-barrel proteins and are indispensable for viability. In Arabidopsis (Arabidopsis thaliana) chloroplasts, there are two distinct types of Omp85-related protein: Toc75 (Translocon at the outer envelope membrane of chloroplasts, 75 kD) and OEP80 (Outer Envelope Protein, 80 kD). Toc75 functions as a preprotein translocation channel during chloroplast import, but the role of OEP80 remains elusive. We characterized three T-DNA mutants of the Arabidopsis OEP80 (AtOEP80) gene. Selectable markers associated with the oep80-1 and oep80-2 insertions segregated abnormally, suggesting embryo lethality of the homozygous genotypes. Indeed, no homozygotes were identified among .100 individuals, and heterozygotes of both mutants produced approximately 25% aborted seeds upon self-pollination. Embryo arrest occurred at a relatively late stage (globular embryo proper) as revealed by analysis using Nomarski optics microscopy. This is substantially later than arrest caused by loss of the principal Toc75 isoform, atToc75-III (two-cell stage), suggesting a more specialized role for AtOEP80. Surprisingly, the oep80-3 T-DNA (located in exon 1 between the first and second ATG codons of the open reading frame) did not cause any detectable developmental defects or affect the size of the AtOEP80 protein in chloroplasts. This indicates that the N-terminal region of AtOEP80 is not essential for the targeting, biogenesis, or functionality of the protein, in contrast with atToc75-III, which requires a bipartite targeting sequence.

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Origin of a chloroplast protein importer

Cited by 144 publications

References 45 publications

Trichomonas vaginalis Hmp35, a Putative Pore-forming Hydrogenosomal Membrane Protein, Can Form a Complex in Yeast Mitochondria

Trichomonas vaginalis Hmp35, a Putative Pore-forming Hydrogenosomal Membrane Protein, Can Form a Complex in Yeast Mitochondria

Evidence for conservation of architecture and physical properties of Omp85-like proteins throughout evolution

The Omp85-Related Chloroplast Outer Envelope Protein OEP80 Is Essential for Viability in Arabidopsis

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