Homologue replacement in the import motor of the mitochondrial inner membrane of trypanosomes
Abstract
Many mitochondrial proteins contain N-terminal presequences that direct them to the organelle. The main driving force for their translocation across the inner membrane is provided by the presequence translocase-associated motor (PAM) which contains the J-protein Pam18. Here, we show that in the PAM of Trypanosoma brucei the function of Pam18 has been replaced by the non-orthologous euglenozoan-specific J-protein TbPam27. TbPam27 is specifically required for the import of mitochondrial presequence-containing but not for carrier proteins. Similar to yeast Pam18, TbPam27 requires an intact J-domain to function. Surprisingly, T. brucei still contains a bona fide Pam18 orthologue that, while essential for normal growth, is not involved in protein import. Thus, during evolution of kinetoplastids, Pam18 has been replaced by TbPam27. We propose that this replacement is linked to the transition from two ancestral and functionally distinct TIM complexes, found in most eukaryotes, to the single bifunctional TIM complex present in trypanosomes.
Data availability
All produced data are contained within the manuscript (e.g. Data Source files)
Article and author information
Author details
Funding
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (175563)
- Andre Schneider
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (NCCR RNA and Disease)
- Andre Schneider
ERC Consolidator grant (648235)
- Bettina Warscheid
Deutsche Forschungsgemeinschaft (403222702/SFB 1381)
- Bettina Warscheid
Germany's Excellence Strategy (CIBSS - EXC-2189 - Project ID 390939984)
- Bettina Warscheid
Excellence Initiative of the German Federal and State Governments (EXC 294 BIOSS)
- Bettina Warscheid
Peter und Traudl Engelhorn foundation
- Anke Harsman
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2020, von Känel et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 1,293
- views
-
- 192
- downloads
-
- 21
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Further reading
-
- Biochemistry and Chemical Biology
- Computational and Systems Biology
The spike protein is essential to the SARS-CoV-2 virus life cycle, facilitating virus entry and mediating viral-host membrane fusion. The spike contains a fatty acid (FA) binding site between every two neighbouring receptor-binding domains. This site is coupled to key regions in the protein, but the impact of glycans on these allosteric effects has not been investigated. Using dynamical nonequilibrium molecular dynamics (D-NEMD) simulations, we explore the allosteric effects of the FA site in the fully glycosylated spike of the SARS-CoV-2 ancestral variant. Our results identify the allosteric networks connecting the FA site to functionally important regions in the protein, including the receptor-binding motif, an antigenic supersite in the N-terminal domain, the fusion peptide region, and another allosteric site known to bind heme and biliverdin. The networks identified here highlight the complexity of the allosteric modulation in this protein and reveal a striking and unexpected link between different allosteric sites. Comparison of the FA site connections from D-NEMD in the glycosylated and non-glycosylated spike revealed that glycans do not qualitatively change the internal allosteric pathways but can facilitate the transmission of the structural changes within and between subunits.
-
- Biochemistry and Chemical Biology
- Genetics and Genomics
Deep Mutational Scanning (DMS) is an emerging method to systematically test the functional consequences of thousands of sequence changes to a protein target in a single experiment. Because of its utility in interpreting both human variant effects and protein structure-function relationships, it holds substantial promise to improve drug discovery and clinical development. However, applications in this domain require improved experimental and analytical methods. To address this need, we report novel DMS methods to precisely and quantitatively interrogate disease-relevant mechanisms, protein-ligand interactions, and assess predicted response to drug treatment. Using these methods, we performed a DMS of the melanocortin-4 receptor (MC4R), a G-protein-coupled receptor (GPCR) implicated in obesity and an active target of drug development efforts. We assessed the effects of >6600 single amino acid substitutions on MC4R’s function across 18 distinct experimental conditions, resulting in >20 million unique measurements. From this, we identified variants that have unique effects on MC4R-mediated Gαs- and Gαq-signaling pathways, which could be used to design drugs that selectively bias MC4R’s activity. We also identified pathogenic variants that are likely amenable to a corrector therapy. Finally, we functionally characterized structural relationships that distinguish the binding of peptide versus small molecule ligands, which could guide compound optimization. Collectively, these results demonstrate that DMS is a powerful method to empower drug discovery and development.