Nishita K, Ojima T, Takahashi A, Inoue A
Department of Chemistry, Faculty of Fisheries, Hokkaido University, Hakodate.
J Biochem. 1997 Mar;121(3):419-24. doi: 10.1093/oxfordjournals.jbchem.a021605.
Troponin which can confer Ca2+-sensitivity upon rabbit actomyosin Mg-ATPase activity has been prepared from the smooth adductor muscle of Ezo-giant scallop (Patinopecten yessoensis). The troponin comprises 40-, 20-, and 19-kDa components. In order to characterize the components, they were separated from each other by CM-Toyopearl column chromatography in the presence of 6 M urea. Consequently, the 20-kDa component was identified as troponin C, based on the Ca2+-binding ability. The amount of Ca2+ bound to the troponin C was estimated to be 0.75 mol/mol at 10(-4) M Ca2+ by the equilibrium dialysis method. The 19-kDa component was identified as troponin I on the basis of not only its inhibitory effect on rabbit actomyosin Mg-ATPase activity along with the smooth adductor tropomyosin, but also the releasing effect of the smooth adductor troponin C on the inhibition. On the other hand, the 40-kDa component was regarded as troponin T on the basis that it bound to F-actin-tropomyosin filament and was indispensable for conferring Ca2+-sensitivity upon rabbit actomyosin Mg-ATPase activity, along with troponin C and troponin I. The above assignments were confirmed by both amino acid analysis and immunoblotting using rabbit antisera raised against counterparts of scallop striated muscle troponin.
已经从北海道巨扇贝（虾夷扇贝）的平滑肌中制备出了肌钙蛋白，它能够赋予兔肌动球蛋白Mg-ATP酶活性以Ca2+敏感性。该肌钙蛋白由40 kDa、20 kDa和19 kDa的组分构成。为了对这些组分进行表征，在6 M尿素存在的情况下，通过CM- Toyopearl柱色谱法将它们彼此分离。结果，基于Ca2+结合能力，将20 kDa的组分鉴定为肌钙蛋白C。通过平衡透析法估计，在10(-4) M Ca2+时，与肌钙蛋白C结合的Ca2+量为0.75 mol/mol。基于其与平滑肌原肌球蛋白一起对兔肌动球蛋白Mg-ATP酶活性的抑制作用，以及平滑肌肌钙蛋白C对该抑制作用的解除效应，将19 kDa的组分鉴定为肌钙蛋白I。另一方面，基于它与F-肌动蛋白-原肌球蛋白丝结合，并且与肌钙蛋白C和肌钙蛋白I一起对于赋予兔肌动球蛋白Mg-ATP酶活性以Ca2+敏感性是不可或缺的，将40 kDa的组分视为肌钙蛋白T。上述归属通过氨基酸分析以及使用针对扇贝横纹肌肌钙蛋白对应物产生的兔抗血清进行的免疫印迹得以证实。
