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Redox Biol. 2018 May;15:363-374. doi: 10.1016/j.redox.2017.12.010. Epub 2017 Dec 23.

A single-cysteine mutant and chimeras of essential Leishmania Erv can complement the loss of Erv1 but not of Mia40 in yeast.

Author information

1
Department of Parasitology, Ruprecht-Karls University, D-69120 Heidelberg, Germany.
2
i3S - Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Porto 4200, Portugal; Institute for Molecular and Cell Biology - IBMC, Universidade do Porto, 4200-135 Porto, Portugal.
3
Biomedical Center Munich - Physiological Chemistry, Ludwig-Maximilians University, D-82152 Planegg, Martinsried, Germany.
4
i3S - Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Porto 4200, Portugal; Institute for Molecular and Cell Biology - IBMC, Universidade do Porto, 4200-135 Porto, Portugal; ICBAS - Instituto de Ciências Biomédicas Abel Salazar, Universidade do Porto, Porto 4050-313, Portugal.
5
Biomedical Center Munich - Physiological Chemistry, Ludwig-Maximilians University, D-82152 Planegg, Martinsried, Germany. Electronic address: kai.hell@bmc.med.lmu.de.
6
Department of Parasitology, Ruprecht-Karls University, D-69120 Heidelberg, Germany; Faculty of Chemistry - Biochemistry, Kaiserslautern University, D-67663 Kaiserslautern, Germany. Electronic address: deponte@chemie.uni-kl.de.

Abstract

Mia40/CHCHD4 and Erv1/ALR are essential for oxidative protein folding in the mitochondrial intermembrane space of yeast and mammals. In contrast, many protists, including important apicomplexan and kinetoplastid parasites, lack Mia40. Furthermore, the Erv homolog of the model parasite Leishmania tarentolae (LtErv) was shown to be incompatible with Saccharomyces cerevisiae Mia40 (ScMia40). Here we addressed structure-function relationships of ScErv1 and LtErv as well as their compatibility with the oxidative protein folding system in yeast using chimeric, truncated, and mutant Erv constructs. Chimeras between the N-terminal arm of ScErv1 and a variety of truncated LtErv constructs were able to rescue yeast cells that lack ScErv1. Yeast cells were also viable when only a single cysteine residue was replaced in LtErvC17S. Thus, the presence and position of the C-terminal arm and the kinetoplastida-specific second (KISS) domain of LtErv did not interfere with its functionality in the yeast system, whereas a relatively conserved cysteine residue before the flavodomain rendered LtErv incompatible with ScMia40. The question whether parasite Erv homologs might also exert the function of Mia40 was addressed in another set of complementation assays. However, neither the KISS domain nor other truncated or mutant LtErv constructs were able to rescue yeast cells that lack ScMia40. The general relevance of Erv and its candidate substrate small Tim1 was analyzed for the related parasite L. infantum. Repeated unsuccessful knockout attempts suggest that both genes are essential in this human pathogen and underline the potential of mitochondrial protein import pathways for future intervention strategies.

KEYWORDS:

ALR; CHCHD4; Erv; Intermembrane space; Leishmania; Mia40; Mitochondria; Oxidative protein folding

PMID:
29310075
PMCID:
PMC5760468
DOI:
10.1016/j.redox.2017.12.010
[Indexed for MEDLINE]
Free PMC Article

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