serval:BIB_438639CA1417
The CcmC-CcmE interaction during cytochrome c maturation by System I is driven by protein-protein and not protein-heme contacts.
10.1074/jbc.RA118.005024
000448515100019
30206118
Shevket
S.H.
author
Gonzalez
D.
author
Cartwright
J.L.
author
Kleanthous
C.
author
Ferguson
S.J.
author
Redfield
C.
author
Mavridou
DAI
author
article
2018-10-26
The Journal of biological chemistry
1083-351X
0021-9258
journal
293
43
16778-16790
Cytochromes c are ubiquitous proteins, essential for life in most organisms. Their distinctive characteristic is the covalent attachment of heme to their polypeptide chain. This post-translational modification is performed by a dedicated protein system, which in many Gram-negative bacteria and plant mitochondria is a nine-protein apparatus (CcmA-I) called System I. Despite decades of study, mechanistic understanding of the protein-protein interactions in this highly complex maturation machinery is still lacking. Here, we focused on the interaction of CcmC, the protein that sources the heme cofactor, with CcmE, the pivotal component of System I responsible for the transfer of the heme to the apocytochrome. Using in silico analyses, we identified a putative interaction site between these two proteins (residues Asp <sup>47</sup> , Gln <sup>50</sup> , and Arg <sup>55</sup> on CcmC; Arg <sup>73</sup> , Asp <sup>101</sup> , and Glu <sup>105</sup> on CcmE), and we validated our findings by in vivo experiments in Escherichia coli Moreover, employing NMR spectroscopy, we examined whether a heme-binding site on CcmE contributes to this interaction and found that CcmC and CcmE associate via protein-protein rather than protein-heme contacts. The combination of in vivo site-directed mutagenesis studies and high-resolution structural techniques enabled us to determine at the residue level the mechanism for the formation of one of the key protein complexes for cytochrome c maturation by System I.
Amino Acid Substitution
Apoproteins/chemistry
Apoproteins/genetics
Apoproteins/metabolism
Bacterial Outer Membrane Proteins/chemistry
Bacterial Outer Membrane Proteins/genetics
Bacterial Outer Membrane Proteins/metabolism
Binding Sites
Crystallography, X-Ray
Cytochromes c/chemistry
Cytochromes c/genetics
Cytochromes c/metabolism
Escherichia coli/genetics
Escherichia coli/growth & development
Escherichia coli/metabolism
Escherichia coli Proteins/chemistry
Escherichia coli Proteins/genetics
Escherichia coli Proteins/metabolism
Heme/chemistry
Heme/genetics
Heme/metabolism
Hemeproteins/chemistry
Hemeproteins/genetics
Hemeproteins/metabolism
Membrane Proteins/chemistry
Membrane Proteins/genetics
Membrane Proteins/metabolism
Mutagenesis, Site-Directed
Protein Conformation
Protein Interaction Domains and Motifs
CcmC
CcmE
Gram-negative bacteria
System I
cytochrome c
cytochrome c maturation
heme
nuclear magnetic resonance (NMR)
post-translational modification (PTM)
protein-protein interactions
eng
60_published
true
peer-reviewed
Publication types: Journal Article ; Research Support, Non-U.S. Gov't
Publication Status: ppublish
University of Lausanne
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