Biochemical evidence for conformational changes in the cross-talk between adenylation and peptidyl-carrier protein domains of nonribosomal peptide synthetases.
Nonribosomal peptide synthetases serve as multidomain protein templates for producing a wealth of pharmaceutically important natural products. For the correct assembly of the desired natural product the interactions between the different catalytic centres and the reaction intermediates bound to the peptidyl carrier protein must be precisely controlled at spatial and temporal levels. We have investigated the interplay between the adenylation (A) domain and the peptidyl carrier protein in the gramicidin S synthetase I (EC 5.1.1.11) via partial tryptic digests, native PAGE and gel-filtration analysis, as well as by chemical labeling experiments. Our data imply that the 4'-phosphopantetheine moiety of the peptidyl carrier protein changes its position as a result of a conformational change in the A domain, which is induced by the binding of an amino acyl adenylate mimic. The productive interaction between the two domains at the stage of the amino acyl transfer onto the 4'-phosphopantetheine moiety is accompanied by a highly compact protein conformation of the holo-protein. These results provide the first biochemical evidence for the occurrence of conformational changes in the cross-talk between A and peptidyl carrier protein domains of a multidomain nonribosomal peptide synthetase.
Zettler
Joachim
J
Technische Universität Dortmund, Dortmund, Germany.
Mootz
Henning D
HD
D000591:Amino Acid Isomerases / Q000737:chemistry*; D002352:Carrier Proteins / Q000737:chemistry*; D002384:Catalysis; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D008958:Models, Molecular; D018716:Molecular Mimicry; D015394:Molecular Structure; D010453:Peptide Synthases / Q000737:chemistry*; D010957:Plasmids / Q000235:genetics; D026723:Polyadenylation; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D002352:Carrier Proteins; D000591:Amino Acid Isomerases; C022343:phenylalanine racemase (ATP-hydrolyzing); D010453:Peptide Synthases; C104385:non-ribosomal peptide synthase