Mutational analysis and homology modelling of SyrC, the aminoacyltransferase in the biosynthesis of syringomycin.
SyrC, a component of the multienzyme system of syringomycin biosynthesis, has been shown to shuttle Thr/4-Cl-Thr between the thiolation domains SyrB1-T1 and SyrE-T8,9 by transiently linking it to Cys224 in the enzyme active site. We present data on the structure-function relationship in vivo of this protein and an in silico model of its three-dimensional structure. The biosynthetic activity of SyrC was not influenced when either Asp348 or His376 that together with Cys224 form a putative catalytic triad, were replaced with Ala, but it was abolished by the exchange Cys224 with Ser. The presence of the FLAG peptide on either the N- or C-terminus of the protein did not affect activity, whereas the deletion of the first 16 amino acids at the N-terminus or the insertion of Maltose Binding Protein abolished the production of syringomycin. We present the model of the three-dimensional structure of SyrC suggesting a homodimeric structure for the protein and biochemical data that are supportive of this model.
Fullone
Maria Rosaria
MR
Department of Biochemical Sciences A. Rossi Fanelli, University of Rome La Sapienza, P.le A. Moro, 5, 00185 Rome, Italy.
Paiardini
Alessandro
A
Gross
Dennis C
DC
Lu
Shi-En
SE
Fiore
Alberto
A
Grgurina
Ingeborg
I
D000595:Amino Acid Sequence; D019881:Aminoacyltransferases / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D002352:Carrier Proteins / Q000235:genetics; D019281:Dimerization; D058619:Maltose-Binding Proteins; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009842:Oligopeptides; D010453:Peptide Synthases / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D010455:Peptides / Q000235:genetics; D011487:Protein Conformation; D044224:Pseudomonas syringae / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D002352:Carrier Proteins; D058619:Maltose-Binding Proteins; D009842:Oligopeptides; D010455:Peptides; D011993:Recombinant Fusion Proteins; C032677:syringomycin; C085706:FLAG peptide; D019881:Aminoacyltransferases; D010453:Peptide Synthases; C094582:SyrC protein, Pseudomonas syringae