期刊
ELECTROPHORESIS
卷 26, 期 1, 页码 47-52出版社
WILEY
DOI: 10.1002/elps.200406148
关键词
biotin-4-fluorescein; functional screening; nondenaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresis; streptavidin
The biotin-binding protein streptavidin exhibits a high stability against thermal denaturation, especially when complexed to biotin, Herein we show that, in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SIDS-PAGE), streptavidin is stabilized at high temperature in the presence of biotinylated fluorescent probes, such as biotin-4-fluorescein, which is incorporated within the binding pocket. In nondenaturing SIDS-PAGE, streptavidin is detectable when complexed with biotin-4-fluorescein using a UV-transilluminator. Using biotin-4-fluorescein, the detection limit of streptavidin lies in the same range as with Coomassie blue staining. The functionality of streptavidin mutants can readily be assessed from crude bacteria] extracts using biotin-4-fluorescein as a probe in nondenaturing SDS-PAGE.
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