Selective labeling of functional groups on membrane proteins or glycoproteins using reactive biotin derivatives and 125I-streptavidin.
Amino groups, sulfhydryl groups or oxidation-induced aldehydes on erythrocyte membrane proteins and/or glycoproteins, were reacted with biotinyl N-hydroxysuccinimide ester (BNHS), 3-(N-maleimido-propionyl) biocytin (MPB) or biocytin hydrazide (BCHZ), respectively. The detergent-lysed biotinylated samples were subjected to SDS-polyacrylamide gel electrophoresis, and the proteins were transferred onto nitrocellulose membranes. The blot was then incubated with a solution containing 125I-streptavidin, and processed for autoradiography. The advantages of this approach over previously reported procedures for labeling the three functional groups include the following: extremely high sensitivity; short exposure times of autoradiograms and relatively low levels of radioactivity; single-step radiolabeling procedures subsequent to processing and handling of gels and no background labeling in control samples.
D001426:Bacterial Proteins / Q000378:metabolism*; D001710:Biotin / Q000031:analogs & derivatives* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004910:Erythrocyte Membrane / Q000378:metabolism; D005470:Fluorometry; D006801:Humans; D006834:Hydrazines / Q000378:metabolism; D008239:Lysine / Q000031:analogs & derivatives / Q000378:metabolism; D008301:Maleimides / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D019809:Streptavidin; D013388:Succinimides
D001426:Bacterial Proteins; D006834:Hydrazines; D008301:Maleimides; D008565:Membrane Proteins; D013388:Succinimides; C033788:biotinyl N-hydroxysuccinimide ester; C048725:biocytin hydrazide; D001710:Biotin; D019809:Streptavidin; C047272:3-(N-maleimidopropionyl)biocytin; D008239:Lysine