National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
R01GM120996
米国
National Institutes of Health/National Cancer Institute (NIH/NCI)
R01CA208834
米国
引用
ジャーナル: Nat Chem Biol / 年: 2023 タイトル: Structural basis of colibactin activation by the ClbP peptidase. 著者: José A Velilla / Matthew R Volpe / Grace E Kenney / Richard M Walsh / Emily P Balskus / Rachelle Gaudet / 要旨: Colibactin, a DNA cross-linking agent produced by gut bacteria, is implicated in colorectal cancer. Its biosynthesis uses a prodrug resistance mechanism: a non-toxic precursor assembled in the ...Colibactin, a DNA cross-linking agent produced by gut bacteria, is implicated in colorectal cancer. Its biosynthesis uses a prodrug resistance mechanism: a non-toxic precursor assembled in the cytoplasm is activated after export to the periplasm. This activation is mediated by ClbP, an inner-membrane peptidase with an N-terminal periplasmic catalytic domain and a C-terminal three-helix transmembrane domain. Although the transmembrane domain is required for colibactin activation, its role in catalysis is unclear. Our structure of full-length ClbP bound to a product analog reveals an interdomain interface important for substrate binding and enzyme stability and interactions that explain the selectivity of ClbP for the N-acyl-D-asparagine prodrug motif. Based on structural and biochemical evidence, we propose that ClbP dimerizes to form an extended substrate-binding site that can accommodate a pseudodimeric precolibactin with its two terminal prodrug motifs in the two ClbP active sites, thus enabling the coordinated activation of both electrophilic warheads.
根拠: light scattering, SEC-MALS performed on protein solubilized in DDM. Conjugate analysis suggests the average protein molecular weight of the particles in the ClbP peak is approx. 110kDa, which ...根拠: light scattering, SEC-MALS performed on protein solubilized in DDM. Conjugate analysis suggests the average protein molecular weight of the particles in the ClbP peak is approx. 110kDa, which matches the molecular weight of the dimer.