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Rabbit fast skeletal muscle phospholipase C Molecular weight determination by renaturation after polyacrylamide gel electrophoresis in the presence of sodium dodecylsulfate
Author(s) -
Windhofer Volker,
Varsànyi Magdolna,
Heilmeyer Ludwig M.G.
Publication year - 1992
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(92)81182-l
Subject(s) - chemistry , sodium , size exclusion chromatography , skeletal muscle , phospholipase c , polyacrylamide gel electrophoresis , enzyme , biochemistry , molecular mass , hydrolysis , chromatography , gel electrophoresis , biology , endocrinology , organic chemistry
Phosphoinositide specific phospholipase C from rabbit fast skeletal muscle has been enriched ca. 1,000‐fold with a specific activity of 40 μmol × min −1 × mg −1 . Following SDS‐PAGE, renaturation of the enzyme protein in the presence of deoxycholate allowed the determination of an apparent molecular weight of 110 kDa. Gel‐filtration of the native enzyme resulted in a very similar apparent molecular weight of 115 kDa, however, associated proteins of higher molecular weight were also found. Free Ca 2+ concentrations needed for half‐maximal activation of PtdIns(4,5)P 2 , PtdIns4P and PtdIns hydrolysis are 6.3 μM, 85 μM and 1.8 mM, and the K m values for these substrates 102, 340 and 937 μM, respectively.