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1.
PLoS Comput Biol ; 19(8): e1011090, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37549177

RESUMO

Complexome profiling allows large-scale, untargeted, and comprehensive characterization of protein complexes in a biological sample using a combined approach of separating intact protein complexes e.g., by native gel electrophoresis, followed by mass spectrometric analysis of the proteins in the resulting fractions. Over the last decade, its application has resulted in a large collection of complexome profiling datasets. While computational methods have been developed for the analysis of individual datasets, methods for large-scale comparative analysis of complexomes from multiple species are lacking. Here, we present Comparative Clustering (CompaCt), that performs fully automated integrative analysis of complexome profiling data from multiple species, enabling systematic characterization and comparison of complexomes. CompaCt implements a novel method for leveraging orthology in comparative analysis to allow systematic identification of conserved as well as taxon-specific elements of the analyzed complexomes. We applied this method to a collection of 53 complexome profiles spanning the major branches of the eukaryotes. We demonstrate the ability of CompaCt to robustly identify the composition of protein complexes, and show that integrated analysis of multiple datasets improves characterization of complexes from specific complexome profiles when compared to separate analyses. We identified novel candidate interactors and complexes in a number of species from previously analyzed datasets, like the emp24, the V-ATPase and mitochondrial ATP synthase complexes. Lastly, we demonstrate the utility of CompaCt for the automated large-scale characterization of the complexome of the mosquito Anopheles stephensi shedding light on the evolution of metazoan protein complexes. CompaCt is available from https://github.com/cmbi/compact-bio.


Assuntos
Eucariotos , Proteínas , Animais , Análise por Conglomerados , Células Eucarióticas/metabolismo , Espectrometria de Massas/métodos , Proteínas/metabolismo
2.
Life Sci Alliance ; 6(7)2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-37094942

RESUMO

Mitochondrial bc 1 complex from yeast has 10 subunits, but only cytochrome b (Cytb) subunit is encoded in the mitochondrial genome. Cytb has eight transmembrane helices containing two hemes b for electron transfer. Cbp3 and Cbp6 assist Cytb synthesis, and together with Cbp4 induce Cytb hemylation. Subunits Qcr7/Qcr8 participate in the first steps of assembly, and lack of Qcr7 reduces Cytb synthesis through an assembly-feedback mechanism involving Cbp3/Cbp6. Because Qcr7 resides near the Cytb carboxyl region, we wondered whether this region is important for Cytb synthesis/assembly. Although deletion of the Cytb C-region did not abrogate Cytb synthesis, the assembly-feedback regulation was lost, so Cytb synthesis was normal even if Qcr7 was missing. Mutants lacking the Cytb C-terminus were non-respiratory because of the absence of fully assembled bc 1 complex. By performing complexome profiling, we showed the existence of aberrant early-stage subassemblies in the mutant. In this work, we demonstrate that the C-terminal region of Cytb is critical for regulation of Cytb synthesis and bc 1 complex assembly.


Assuntos
Citocromos b , Proteínas de Saccharomyces cerevisiae , Citocromos b/genética , Citocromos b/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Complexo III da Cadeia de Transporte de Elétrons , Saccharomyces cerevisiae/metabolismo , Mitocôndrias/metabolismo , Proteínas de Transporte , Proteínas de Membrana/metabolismo , Chaperonas Moleculares/metabolismo , Proteínas Mitocondriais/genética
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