Chemical modification with dihydro-4,4'-diisothiocyanostilbene-2,2'-disulfonate reveals the distance between K480 and K501 in the ATP-binding domain of the Na,K-ATPase.
Chemical modification with dihydro-4,4'-diisothiocyanostilbene-2,2'-disulfonate reveals the distance between K480 and K501 in the ATP-binding domain of the Na,K-ATPase.
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用二氢-4,4-二异硫氰芪-2,2-二磺酸盐进行化学修饰揭示了 Na,K-ATP 酶 ATP 结合域中 K480 和 K501 之间的距离。
DOI：
10.1006/abbi.1997.9879
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发表时间：
1997
期刊：
影响因子：
--
通讯作者：
Kaplan,JH
中科院分区：
文献类型：
--
作者：
Gatto,C;Lutsenko,S;Kaplan,JH
Dihydro-4,4′-diisothiocyanostilbene-2,2′-disulfonate (H2DIDS) inactivates the renal Na,K-ATPase in an ATP- and K-preventable fashion; inactivation results in the covalent incorporation of a single [3H2]DIDS molecule into the Na pump α-subunit. K+protection is observed at low concentrations (<2 mm) and reversed at higher concentrations. The biphasic effect is also seen with Rb+, to a lesser extent by Cs+, and not at all by Na+or choline. After extensive tryptic digestion of3H2DIDS-inactivated enzyme, a single radiolabeled peptide is seen in 16.5% Tricine gels. N-terminal amino acid sequencing revealed two sequences470IVEIPFNSTNxYQLS and495HLLVMxGAPER, the unidentified residues were K480and K501, respectively. These data provide suggestive evidence of cross-linking by H2DIDS between the two lysines. CNBr digestion of3H2DIDS-labeled α-subunit produced a single radioactive band of the predicted 15-kDa mass for cross-linking between K480an K501produced by cleavage at known methione residues. The 15-kDa band combined two N-terminal sequences464RDRYAKIVEI and501xGAPERILDR which include K480and K501. Thus K480and K501are within approximately 14 Å of each other in the Na-bound form of the enzyme and information about the occupancy of the cation binding domain is transmitted to the ATP binding loop of the Na,K-ATPase.
