In this Account, we examine the characteristics of the enzymes responsible for constructing AviCys to evaluate possibilities for generating high yields of bioactive AviCys- or AviMeCys-containing peptides for research and clinical
use.\n\nThe gene cluster for the biosynthesis of epidermin has been studied in depth, leading to the proposal for Barasertib Cell Cycle inhibitor a mechanism of AviCys formation. First, a serine residue upstream of the C-terminus is enzymatically dehydrated to form a dehydroalanine residue. Then, the C-terminal cysteine residue is oxidatively decarboxylated to form an enethiolate, which subsequently cyclizes onto the dehydroalanine to give the AviCys ring. Extensive research on EpiD, the enzyme responsible for the oxidative decarboxylation reaction, has led to its purification and cocrystallization with a model substrate peptide, yielding an X-ray crystal structure. An in vitro assay of the enzyme with a library of synthetic heptapeptides has resulted in the discovery that EpiD has low absolute substrate specificity and can oxidatively decarboxylate a wide variety of C-terminal cysteine-containing peptides.\n\nRecently, the gene duster for the biosynthesis of cypemycin
was also identified. Despite certain structural similarities between cypemycin and the lantibiotic peptides, analysis of the biosynthetic genes suggests that cypemycin ABT-263 mouse production is quite different from that of the lantibiotics. In particular, the AviCys residue in cypemycin is formed from two cysteine Napabucasin research buy residues instead of one serine and one cysteine, and the CypD enzyme that catalyzes the oxidative decarboxylation of the C-terminal cysteine shows little homology to EpiD.\n\nThe knowledge accrued from studying EpiD and CypD could be used to develop a semisynthetic methodology to produce
AviCys-containing peptides. In particular, suitable precursor peptides could be synthesized on solid support before being fed to either of these enzymes in vitro to generate the C-terminal AviCys moiety. Exploring the potential of this methodology could lead to the efficient production of epidermin, cypemycin, and analogues thereof.”
“The aims of this study were to investigate mechanisms of action involved in H2AX phosphorylation by DNA interstrand crosslinking (ICL) agents and determine whether gamma H2AX could be a suitable pharmacological marker for identifying potential ICL cellular chemosensitivity. In normal human fibroblasts, after treatment with nitrogen mustard (HN2) or cisplatin, the peak gamma H2AX response was detected 2-3 h after the peak of DNA ICLs measured using the comet assay, a validated method for detecting ICLs in vitro or in clinical samples. Detection of gamma H2AX foci by immunofluorescence microscopy could be routinely detected with 6-10 times lower concentrations of both drugs compared to detection of ICLs using the comet assay.