The sulfhydryl groups involved in the active site of myosin B adenosinetriphosphatase. VII. A chemical modification of colonic smooth myosin B with N-ethylmaleimide.
The reactivity of the sulfhydryl groups in colonic myosin B to N-ethylmaleimide (NEM) was studied under various conditions. A higher concentration of NEM was required to alter the ATPase activity as compared with skeletal myosin B. A chemical modification of colonic myosin B at a low ionic strength brought about the marked inhibition of EDTA-ATPase and 30-40% activation of Mg2+-ATPase, but had little effect on Ca2+-ATPase. The same results were obtained with NEM modification at a high ionic strength. The presence of 10 mM MgCl2 in the reaction medium for NEM modification caused the activation of Ca2+-ATPase and remarkable inhibition of EDTA-ATPase, suggesting that SH1 is here reactive to NEM. NEM modification of colonic myosin B in the presence of ATP resulted in the inhibition of not only EDTA-ATPase but also Ca2+- and Mg2+-ATPases although the degree of inhibition was different. Addition of 2.0 M urea to the modification medium caused the simultaneous inhibition of EDTA- and Ca2+-ATPase but little change in Mg2+-ATPase. Based on these results, the presence of three specific sulfhydryl groups, SHa, SH1, and SH2, is discussed.
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D003106:Colon / Q000201:enzymology; D004285:Dogs; D004789:Enzyme Activation / Q000187:drug effects; D005033:Ethylmaleimide / Q000494:pharmacology*; D008274:Magnesium / Q000494:pharmacology; D015636:Magnesium Chloride; D009130:Muscle, Smooth / Q000201:enzymology*; D009132:Muscles / Q000201:enzymology; D009218:Myosins / Q000378:metabolism*; D009994:Osmolar Concentration
D015636:Magnesium Chloride; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D017301:Ca(2+) Mg(2+)-ATPase; C026049:EDTA-ATPase; D009218:Myosins; D000252:Calcium-Transporting ATPases; D008274:Magnesium; D005033:Ethylmaleimide