Cu-free cycloaddition for identifying catalytic active adenylation domains of nonribosomal peptide synthetases by phage display.
To engineer the substrate specificities of nonribosomal peptide synthetases (NRPS), we developed a method to display NRPS modules on M13 phages and select catalytically active adenylation (A) domains that would load azide functionalized substrate analogs to the neighboring peptidyl carrier protein (PCP) domains. Biotin conjugated difluorinated cyclooctyne was used for copper free cycloaddition with an azide substituted substrate attached to PCP. Biotin-labeled phages were selected by binding to streptavidin.
Zou
Yekui
Y
Department of Chemistry, The University of Chicago, 929 E. 57th Street, GCIS, E505A, Chicago, IL 60637, USA.
Yin
Jun
J
D001386:Azides / Q000737:chemistry; D017104:Bacteriophage M13 / Q000737:chemistry*; D001710:Biotin / Q000737:chemistry*; D002384:Catalysis; D020134:Catalytic Domain; D002626:Chemistry, Pharmaceutical / Q000379:methods; D003300:Copper / Q000737:chemistry*; D015195:Drug Design; D008956:Models, Chemical; D019151:Peptide Library; D010453:Peptide Synthases / Q000737:chemistry*; D019809:Streptavidin / Q000737:chemistry*; D013379:Substrate Specificity
D001386:Azides; D019151:Peptide Library; D001710:Biotin; D003300:Copper; D019809:Streptavidin; D010453:Peptide Synthases