MOLECULAR WEIGHT DETERMINATION AND METAL ION REQUIREMENT OF PHOSPHATIDATE PHOSPHOHYDROLASE PURIFIED FROM CYTOSOLIC FRACTION OF RAT LIVER
نویسندگان: ثبت نشده
چکیده مقاله:
Phosphatidate phosphohydrolase (PAP) from cytosolic fraction of rat liver was purified to homogeneity having specific activity of 5.14 U/mg protein. An activity staining procedure was developed to determine molecular weight of the enzyme on polyacrylamide gel electrophoresis using Ferguson plot. Molecular Weight (M.W.) of the active PAP was 298 KDa. SDS-PAGE analysis showed a M.W. of 47 KDa for PAP subunits. Active multimer of the enzyme, therefore, was calculated to be hexamer. Gel filtration on Sephadex G-100 column showed a M.W. of 850 KDa for PAP due to the protein aggregation on the matrix. The purified enzyme was inhibited by divalent cations such as Fe2+, Cu2+ and Ca2+ but requires Mg 2+ for its activity. The activity loss of PAP inhibited by cations was restored by Mg2+ on polyacrylamide gel. The data suggest that the active form of cytosolic PAP is a hexamer of identical subunits and that charge density plays an important role in enzyme-substrate interaction. Magnesium ion is probably the only divalent cation capable of generating proper enzyme-metal-substrate complex necessary for the catalytic activity.
منابع مشابه
molecular weight determination and metal ion requirement of phosphatidate phosphohydrolase purified from cytosolic fraction of rat liver
phosphatidate phosphohydrolase (pap) from cytosolic fraction of rat liver was purified to homogeneity having specific activity of 5.14 u/mg protein. an activity staining procedure was developed to determine molecular weight of the enzyme on polyacrylamide gel electrophoresis using ferguson plot. molecular weight (m.w.) of the active pap was 298 kda. sds-page analysis showed a m.w. of 47 kda for...
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آنزیم تریپسین در شرایط قلیایی ناپایدار می باشد .و فعالیت پروتئولیتیکی تریپسین منجربه خود هضمی آن در جایگاههای خاصی می گردد. بنابر این آنزیمی با ناپایداری بالا محسوب میگردد. در سالهای اخیر موفق شدند که با ایجاد تغیرات شیمیایی با اضافه کردن فلزات خاص ، کلسیم و یا عمل استیلاسیون منجر به افزایش پایداری آنزیم تریپسین گردند. مطالعات در حال حاضر نشان می دهد که تریپسین استیله شده فعالیت آنزیمی خود را ...
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عنوان ژورنال
دوره 13 شماره 3
صفحات -
تاریخ انتشار 2002-09-01
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