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The purified rabbit muscle creatine kinase (R-CK) was previously considered homogeneousand without disulfide bonds.By the method of NR/R two-dimensional diagonal SDS-PAGE,two forms of R-CK,designated respectively “oxidized form” of creatine kinase which contained intrachain disulfide bondsand “reduced form” of creatine kinase which did not have any —S—S— bridges,were for the first time sepa-rated.They were found to be the same in amino acid composition,in subunit molecular Weight and in isoelec-tric point,and were almost identical in enzyme activities.Thus it is hard to isolate one from the other bycommon biochemical methods.More extensive studies show that the oxidized form of CK also contains a pair of reactive thiol groupswhich are essential to the enzyme activity,and it has one intrachain disulfide bond per subunit.In the nativestate,this —S—S— bond cannot be reduced by DTT,but by treating the reduced form of CK with some ox-idants,these —S—S— bonds can be formed in vitro.Thus it is presumed that the disulfide bonds are cross-linked through the oxidization of two shallowly buried —SH groups.
The purified rabbit muscle creatine kinase (R-CK) was previously considered homogeneousand without disulfide bonds.By the method of NR / R two-dimensional diagonal SDS-PAGE, two forms of R-CK, designated respectively “oxidized form” of creatine kinase which contained intrachain disulfide bonds and “reduced form ” of creatine kinase which did not have any -S-S-bridges, were for the first time sepa-rated. The were found to be the same in amino acid composition, in subunit molecular weight and in isoelec-tric point, and were almost identical in enzyme activities. It is hard to isolate one from the other bycommon biochemical methods. Million extensive studies show that the oxidized form of CK also contains a pair of reactive thiol groupswhich are essential to the enzyme activity, and it has one intrachain disulfide bond per subunit. in the native state, this -S-S-bond can not be reduced by DTT, but by treating the reduced form of CK with some ox- idants, these- S-S-bonds can be for med in vitro.Thus it is presumed that the disulfide bonds are cross-linked through the oxidization of two shallowly buried -SH groups.