Defining the "fast-reacting" thiols of myosin by reaction with 1, 5 IAEDANS. |
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Authors: | R Takashi J Duke K Ue M F Morales |
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Affiliation: | Cardiovascular Research Institute, University of California, San Francisco, California 94143 U.S.A. |
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Abstract: | The specificity of the fluorescent reagent N-iodoacetyl-N-(5-sulfo-1-naphthyl)ethylenediamine (1,5 IAEDANS) for a specific thiol group of myosin has been characterized by a comparison with iodoacetamide (IAA) and by observing maximal enhancement of the Ca2+-ATPase activity and inhibition of the K+-EDTA-ATPase activity of myosin. The stoichiometry of the [3H]1,5 IAEDANS bound to myosin indicates the presence of two fast-reacting thiols which correspond to the “SH1” groups responsible for the catalytic properties of myosin. Moreover, it has been unequivocally demonstrated by gel electrophoresis that the fast-reacting thiol is located on the myosin heavy chain. A single radioactivity-labeled thiol peptide obtained from tryptic digests of myosin labeled with [3H]1,5 IAEDANS or iodo[1-14C]acetamide indicates strongly that the identical thiol was labeled by both reagents. |
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