Berrebi-Bertrand I, Maixent J M, Guede F G, Gerbi A, Charlemagne D, Lelievre L G
Laboratoire de Pharmacologie des Transports Ioniques Membranaires, Université Paris VII, France.
Eur J Biochem. 1991 Feb 26;196(1):129-33. doi: 10.1111/j.1432-1033.1991.tb15795.x.
Guinea pig left ventricular muscle contains two distinct molecular forms of the Na+/K(+)-ATPase catalytic alpha subunit. Sarcolemmal vesicles highly enriched in Na+/K(+)-ATPase were isolated by a new procedure that yielded specific activities of 60-100 mumol Pi.h-1.mg-1. SDS/PAGE of isolated sarcolemma after reduction and alkylation of the sulfhydryl groups and identification on immunoblots with specific anti-(alpha subunit) antibodies indicated the presence of two major polypeptides of 100 kDa and 103 kDa, respectively. The two alpha subunits were functional: the dose/response curves of Na+/K(+)-ATPase activity with ouabain, dihydroouabain and digitoxigenin were biphasic, revealing the presence of high-affinity [concentration of drug causing 50% inhibition (IC50) = 10 nM] and low-affinity (IC50 = 2 microM) forms with proportional contributions of 55% and 45%, respectively. The involvement of the high-affinity form in the positive inotropic effect of digitalis and of the low-affinity sites in both inotropy and toxicity are consistent with the literature data on rodents.
豚鼠左心室肌含有两种不同分子形式的Na⁺/K⁺-ATP酶催化α亚基。通过一种新方法分离出高度富集Na⁺/K⁺-ATP酶的肌膜囊泡，其比活性为60 - 100 μmol Pi·h⁻¹·mg⁻¹。对分离的肌膜进行巯基还原和烷基化处理后进行SDS/PAGE，并使用特异性抗（α亚基）抗体在免疫印迹上进行鉴定，结果表明分别存在100 kDa和103 kDa的两种主要多肽。这两种α亚基具有功能：哇巴因、双氢哇巴因和洋地黄毒苷对Na⁺/K⁺-ATP酶活性的剂量/反应曲线呈双相，表明存在高亲和力[导致50%抑制的药物浓度（IC50）= 10 nM]和低亲和力（IC50 = 2 μM）形式，其比例分别为55%和45%。高亲和力形式参与洋地黄的正性肌力作用，低亲和力位点参与肌力作用和毒性作用，这与啮齿动物的文献数据一致。
