Crystal structure of the peptidyl-cysteine decarboxylase EpiD complexed with a pentapeptide substrate

Abstract

Epidermin from Staphylococcus epidermidis Tu È 3298 is an antimicrobial peptide of the lantibiotic family that contains, amongst other unusual amino acids, S-[(Z)-2-aminovinyl]-D-cysteine. This residue is introduced by post-translational modi®cation of the ribosomally synthesized precursor EpiA. Modi®cation starts with the oxidative decarboxylation of its C-terminal cysteine by the¯avoprotein EpiD generating a reactive (Z)-enethiol intermediate. We have determined the crystal structures of EpiD and EpiD H67N in complex with the substrate pentapeptide DSYTC at 2.5 A Ê resolution. Rossmann-type monomers build up a dodecamer of 23 point symmetry with trimers disposed at the vertices of a tetrahedron. Oligomer formation is essential for binding of¯avin mononucleotide and substrate, which is buried by an otherwise disordered substrate recognition clamp. A pocket for the tyrosine residue of the substrate peptide is formed by an induced ®t mechanism. The substrate contacts¯avin mononucleotide only via Cys-Sg, suggesting its oxidation as the initial step. A thioaldehyde intermediate could undergo spontaneous decarboxylation. The unusual substrate recognition mode and the type of chemical reaction performed provide insight into a novel family of¯avoproteins.

Genome mining and genetic analysis of cypemycin biosynthesis reveal an unusual class of posttranslationally modified peptides

“…In contrast, previously described lantibiotic AviCys residues are generated from an internal serine (Ser) and a C-terminal Cys. LanD-mediated oxidative decarboxylation of the propeptide's C-terminal Cys yields an enethiol intermediate that can form AviCys by addition to a Dha created by dehydration of an internal Ser (11).…”

Functional Characterization of the Yeast Ppz1 Phosphatase Inhibitory Subunit Hal3

“…Those seven residues are clustered in three groups. I446K and W452G are located in the loop connecting ␣5 and ␤6, N478D and I480F are in the NH 2 -terminal moiety of helix ␣6, and E460G, V462A, and N466I are located around 20 Å apart from the other two groups in an area known as the flap (12,31). This area has been shown to be unstructured in AtHal3 and EpiD, unless a substrate is bound to the protein (29,30).…”

4′-Phosphopantetheine Biosynthesis in Archaea

“…The absorbance maximum of the flavin cofactor is at 448 nm for the wt enzyme and at 453 nm for the mutant enzyme H87N. From the published crystal structures of PPC decarboxylases and LanD enzymes (8,10,18,19), we know that the conserved His residue of HFCD (homo-oligomeric flavin-dependent Cys decarboxylases) enzymes is in direct neighborhood of the flavin cofactor. Therefore, it is not surprising that exchanging His 87 led to changes in the UV-visible spectrum of the bound cofactor.…”

“…Although several active site residues of eubacterial and eukaryotic CoaC activities are not conserved in Methanocaldococcus Dfp (12), we were able to demonstrate its PPC decarboxylase activity. Furthermore, it is shown that HisCoaC (and thus Dfp) forms homododecamers like the E. coli Dfp protein and the related peptidyl-cysteine decarboxylases EpiD and MrsD (7,10,13). We show that M. jannaschii CoaB is a CTP-dependent enzyme, although the nucleotide binding motif II contains a Lys residue, which is characteristic for the ATP-dependent PPC synthetases (5).…”

“…Subsequent reduction of 4Ј-phosphopantothenoylaminoenethiol to 4Ј-phosphopantetheine by the reduced cofactor FMNH 2 depends on the conserved cysteine residue of the 16-amino acid 4Ј-phosphopantothenoylcysteine binding clamp, 151 PDSGSQACGDIGPGRM (6,8,9). Binding of 4Ј-phosphopantothenoylcysteine also involves the Asn residue of the PXMNXXMW motif, which contacts the carboxyl group (8,10). The presence of conserved CoA biosynthetic genes indicates that all Archaea convert 4Ј-phosphopantothenate into coenzyme A by using CoaB, CoaC, CoaD, and CoaE activities, although cloning and functional characterization of archaebacterial coenzyme A biosynthetic genes has not been published until now (11).…”