Development of a radioactive protein A-based assay for analysis of surface protein expression in gram-positive bacteria.
This paper describes an immunochemical method which uses radioactive protein A for the detection and analysis of streptococcal M6 protein epitopes on the surface of recombinant Streptococcus gordonii. With this assay, recombinant S. gordonii cells expressing a portion of the M6 protein on their surfaces show a 75-fold increase in bound radioactivity over cells of the control S. gordonii parental strain. Furthermore, use of the assay to monitor the amount of M6 protein present on the surface of the S. gordonii recombinant during growth in culture demonstrated that expression is highest at late log phase, with the protein being sloughed off during stationary phase. This simple assay allows analysis of surface protein without any protein purification or sophisticated instrumentation. As such, it should be broadly applicable to following the expression of most surface-accessible bacterial proteins.
Coyle
T C
TC
Department of Microbiology, Oregon State University, Corvallis, USA.
Franke
C A
CA
Hruby
D E
DE
D000942:Antigens, Bacterial / Q000032:analysis / Q000235:genetics; D000954:Antigens, Surface / Q000032:analysis / Q000235:genetics; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D002352:Carrier Proteins; D000939:Epitopes / Q000032:analysis / Q000235:genetics; D005069:Evaluation Studies as Topic; D006094:Gram-Positive Bacteria / Q000737:chemistry*; D007120:Immunochemistry; D008565:Membrane Proteins / Q000032:analysis / Q000235:genetics / Q000276:immunology; D011863:Radioimmunoassay / Q000379:methods* / Q000706:statistics & numerical data; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity; D013205:Staphylococcal Protein A; D013291:Streptococcus / Q000737:chemistry* / Q000235:genetics / Q000254:growth & development; D013462:Sulfur Radioisotopes
D000942:Antigens, Bacterial; D000954:Antigens, Surface; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; D000939:Epitopes; D008565:Membrane Proteins; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A; D013462:Sulfur Radioisotopes; C014689:streptococcal M protein