BOCILLIN FL, a sensitive and commercially available reagent for detection of penicillin-binding proteins.
We describe a new, sensitive, rapid, and nonradioactive method involving the use of the commercially available BOCILLIN FL, a fluorescent penicillin, as a labeling reagent for the detection and study of penicillin-binding proteins (PBPs). This method allowed rapid detection of 30 ng of a purified PBP protein under UV light and of 2 to 4 ng of the protein with the aid of a FluorImager. This method also allowed rapid determination of the PBP profiles of Escherichia coli, Pseudomonas aeruginosa, and Streptococcus pneumoniae. The PBP profiles obtained are virtually identical to those reported previously with 3H-, 14C-, or 125I-labeled penicillin. Using this method enabled us to determine the 50% inhibitory concentrations of the penicillin-sensitive and -resistant PBP2x proteins of S. pneumoniae for penicillin G, thereby allowing a direct evaluation of their relative affinities for penicillin G. Finally, this method also allowed us to compare relative affinities of a PBP2x protein for different beta-lactam antibiotics with the aid of fluorescence polarization technology and to monitor a PBP2x protein during purification.
Zhao
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Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285-0438, USA. Zhao_Genshi@Lilly.com
Meier
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Kahl
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Gee
K R
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Blaszczak
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D001419:Bacteria / Q000378:metabolism; D001426:Bacterial Proteins; D001681:Biological Assay; D001896:Boron Compounds / Q000378:metabolism; D002352:Carrier Proteins / Q000032:analysis*; D005456:Fluorescent Dyes / Q000378:metabolism; D006602:Hexosyltransferases; D002267:Muramoylpentapeptide Carboxypeptidase / Q000032:analysis*; D046915:Penicillin-Binding Proteins; D010406:Penicillins / Q000378:metabolism; D010458:Peptidyl Transferases; D011485:Protein Binding; D012680:Sensitivity and Specificity
C118961:BOCILLIN FL; D001426:Bacterial Proteins; D001896:Boron Compounds; D002352:Carrier Proteins; D005456:Fluorescent Dyes; D046915:Penicillin-Binding Proteins; D010406:Penicillins; D010458:Peptidyl Transferases; D006602:Hexosyltransferases; D002267:Muramoylpentapeptide Carboxypeptidase