二环己基碳二亚胺和环匹阿尼酸对笼形ATP和笼形Ca2+光解诱导的肌浆网差示傅里叶变换红外光谱的影响。

The effect of dicyclohexylcarbodiimide and cyclopiazonic acid on the difference FTIR spectra of sarcoplasmic reticulum induced by photolysis of caged-ATP and caged-Ca2+.

作者信息

Buchet R, Jona I, Martonosi A

机构信息

Department of Biochemistry and Molecular Biology, State University of New York, Syracuse 13210.

出版信息

Biochim Biophys Acta. 1992 Feb 17;1104(1):207-14. doi: 10.1016/0005-2736(92)90152-c.

DOI:10.1016/0005-2736(92)90152-c

PMID:1532328

Abstract

The photochemical release of Ca2+ from caged-Ca2+ in the absence of ATP, and the release of ATP from caged-ATP in the presence of Ca2+ induce characteristic difference FTIR spectra on rabbit sarcoplasmic reticulum that are related to the formation of Ca2-E1 and E approximately P intermediates of the Ca(2+)-ATPase, respectively. Dicyclohexylcarbodiimide (10 nmol/mg protein) abolished both the Ca(2+)-and ATP-induced difference FTIR spectra parallel with inhibition of ATPase activity. Cyclopiazonic acid (50 nmol/mg protein) inhibited the Ca(2+)-induced difference spectrum measured in the absence of ATP, but had no significant effect on the ATP-induced difference spectrum measured in the presence of 1 mM Ca2+. The dog kidney Na+,K(+)-ATPase did not give significant difference spectrum after photolysis of caged-ATP in Ca(2+)-free media containing 90 mM Na+ and 10 mM K+, with or without ouabain. We propose that both the Ca2+ and the ATP-induced difference FTIR spectra of the Ca(2+)-ATPase reflect the occupancy of the high-affinity Ca2+ transport site of the enzyme.

摘要

在无ATP的情况下，从笼形Ca²⁺光化学释放Ca²⁺，以及在有Ca²⁺存在时从笼形ATP释放ATP，会在兔肌浆网上诱导出特征性的差异傅里叶变换红外光谱，分别与Ca²⁺ - ATP酶的Ca²⁺ - E1和E～P中间体的形成有关。二环己基碳二亚胺（10 nmol/mg蛋白质）消除了Ca²⁺和ATP诱导的差异傅里叶变换红外光谱，同时抑制了ATP酶活性。环匹阿尼酸（50 nmol/mg蛋白质）抑制了在无ATP情况下测得的Ca²⁺诱导的差异光谱，但对在1 mM Ca²⁺存在下测得的ATP诱导的差异光谱没有显著影响。在含有90 mM Na⁺和10 mM K⁺的无Ca²⁺培养基中，无论有无哇巴因，对笼形ATP进行光解后，犬肾Na⁺,K⁺ - ATP酶均未产生显著的差异光谱。我们认为，Ca²⁺ - ATP酶的Ca²⁺和ATP诱导的差异傅里叶变换红外光谱均反映了该酶高亲和力Ca²⁺转运位点的占据情况。