Selective labeling of functional groups on membrane proteins or glycoproteins using reactive biotin derivatives and 125I-streptavidin |
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| Authors: | E Roffman L Meromsky H Ben-Hur E A Bayer M Wilchek |
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| Institution: | 1. Saint Petersburg State University, 7/9 Universitetskaya Nab., 199034 Saint Petersburg, Russian Federation;2. Institute of Macromolecular Compounds of Russian Academy of Sciences, V.O. Bolshoii Pr. 31, 199004 Saint Petersburg, Russian Federation;1. Radiopharmaceuticals Division, Bhabha Atomic Research Centre, Mumbai 400085, India;2. Homi Bhabha National Institute, Anushakti Nagar, Mumbai 400094, India;1. Departamento de Química Inorgánica, Universidade de Vigo, 36310 Vigo, Spain;2. CFisUC, Department of Physics, University of Coimbra, Rua Larga, 3004-516 Coimbra, Portugal;3. Departamento de Genética, Bioquímica e Inmunología, Universidade de Vigo, 36310 Vigo, Spain;4. Instituto de Investigación Sanitaria Galicia Sur, Vigo, Spain;1. School of Chemistry and Chemical Engineering, Southeast University, Nanjing, 211189, China;2. Qinzhou University, 12 Binhai Avenue, Qinzhou, Guangxi, China;3. Affiliated Hospital, Guilin Medical College, 15 Lequn Road, Guilin, Guangxi, China;4. Stanford Cancer Institute, Member of Academic Council, Stanford University, USA |
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| Abstract: | 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. |
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