title: beta-Hydroxylation of the aspartyl residue in the phytotoxin syringomycin E: characterization of two candidate hydroxylases AspH and SyrP in Pseudomonas syringae.
abstract: The pseudomonal phytotoxin syringomycin E and related nonribosomal peptides contain an L- threo-beta-hydroxyaspartyl residue at the eighth position of the lipodepsipeptide backbone as part of a conserved nonproteinogenic tripeptide motif. Informatic analysis of the P. syringae genome suggests only one putative non-heme iron hydroxylase, AspH. On heterologous expression in Escherichia coli AspH shows robust catalytic activity with free L-Asp and L-Asp thioesters to make beta-OH-Asp but yields the erythro diastereomer rather than the threo configuration that is found in syringomycin. Further analysis of the Syr gene cluster indicated that SyrP, previously annotated as the gene regulatory protein for the five-gene Syr cluster, is actually homologous to the known non-heme mononuclear iron hydroxylase TauD. Indeed, purified SyrP acts on Asp tethered as the protein-bound S-pantetheinyl thioester on the eighth module of the SyrE megasynthetase. The hydroxylation gives the anticipated L- threo-3-OH-Asp diastereomer found in syringomycin. The knockout of syrP abolishes the production of the mature syringomycin E, while knockout of aspH has no effect on syringomycin production.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: Gitanjali M
authors.initials: GM
authors.lastname: Singh
authors.forename: Pascal D
authors.initials: PD
authors.lastname: Fortin
authors.forename: Alexander
authors.initials: A
authors.lastname: Koglin
authors.forename: Christopher T
authors.initials: CT
authors.lastname: Walsh
mesh_terms: D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D006900:Hydroxylation; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000378:metabolism; D010456:Peptides, Cyclic / Q000235:genetics* / Q000378:metabolism; D044224:Pseudomonas syringae / Q000235:genetics* / Q000378:metabolism; D014118:Toxins, Biological / Q000235:genetics* / Q000378:metabolism
chemical_list: D010456:Peptides, Cyclic; D014118:Toxins, Biological; C105246:syringomycin E; D006899:Mixed Function Oxygenases