How the MccB bacterial ancestor of ubiquitin E1 initiates biosynthesis of the microcin C7 antibiotic.
Publication Type:Journal Article
Source:EMBO J, Volume 28, Issue 13, p.1953-64 (2009)
Keywords:Amino Acid Sequence, Anti-Bacterial Agents, Aspartic Acid, Bacteriocins, Catalytic Domain, Crystallography, X-Ray, Escherichia coli, Escherichia coli Proteins, Humans, Ligases, Models, Molecular, Molecular Sequence Data, Mutation, Nucleotides, Peptides, Protein Binding, Protein Conformation, Protein Processing, Post-Translational, Sequence Alignment, Ubiquitin-Activating Enzymes
<p>The 39-kDa Escherichia coli enzyme MccB catalyses a remarkable posttranslational modification of the MccA heptapeptide during the biosynthesis of microcin C7 (MccC7), a 'Trojan horse' antibiotic. The approximately 260-residue C-terminal region of MccB is homologous to ubiquitin-like protein (UBL) activating enzyme (E1) adenylation domains. Accordingly, MccB-catalysed C-terminal MccA-acyl-adenylation is reminiscent of the E1-catalysed activation reaction. However, unlike E1 substrates, which are UBLs with a C-terminal di-glycine sequence, MccB's substrate, MccA, is a short peptide with an essential C-terminal Asn. Furthermore, after an intramolecular rearrangement of MccA-acyl-adenylate, MccB catalyses a second, unique reaction, producing a stable phosphoramidate-linked analogue of acyl-adenylated aspartic acid. We report six-crystal structures of MccB in apo, substrate-, intermediate-, and inhibitor-bound forms. Structural and kinetic analyses reveal a novel-peptide clamping mechanism for MccB binding to heptapeptide substrates and a dynamic-active site for catalysing dual adenosine triphosphate-consuming reactions. The results provide insight into how a distinctive member of the E1 superfamily carries out two-step activation for generating the peptidyl-antibiotic MccC7.</p>