用4-乙酰氨基-4'-异硫氰基芪-2,2'-二磺酸（SITS）修饰赖氨酸-501使钠钾ATP酶失活。

Inactivation of the Na,K-ATPase by modification of Lys-501 with 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS).

作者信息

Pedemonte C H, Kirley T L, Treuheit M J, Kaplan J H

机构信息

Department of Physiology, University of Pennsylvania, Philadelphia 19104-6085.

出版信息

FEBS Lett. 1992 Dec 7;314(1):97-100. doi: 10.1016/0014-5793(92)81470-7.

DOI:10.1016/0014-5793(92)81470-7

PMID:1333419

Abstract

The sodium pump or Na,K-ATPase, maintains the Na+ and K+ gradients across eukaryotic cell membranes at the expense of ATP. Incubation of purified canine renal Na,K-ATPase with 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS) inhibited the ATPase activity. Both the labeling of the protein and the loss of ATPase activity were prevented by co-incubation with ADP (acting as an ATP analog) or KCl. Only the alpha-subunit was labeled by SITS. The alpha-subunit from the inhibited enzyme was extensively digested with trypsin, and SITS-labeled peptides were purified by reverse-phase HPLC and sequenced. The amino acid sequence determined, His-Leu-Leu-Val-Met-X-Gly-Ala-Pro-Glu, indicated that SITS modifies Lys-501 (X) on the alpha-subunit of Na,K-ATPase.

摘要

钠泵或钠钾 - ATP酶，以ATP为代价维持真核细胞膜上的Na⁺和K⁺梯度。用4 - 乙酰氨基 - 4'-异硫氰基芪 - 2,2'-二磺酸（SITS）孵育纯化的犬肾钠钾 - ATP酶会抑制ATP酶活性。与ADP（作为ATP类似物）或KCl共同孵育可防止蛋白质的标记和ATP酶活性的丧失。只有α亚基被SITS标记。用胰蛋白酶对受抑制酶的α亚基进行广泛消化，通过反相高效液相色谱法纯化SITS标记的肽并进行测序。确定的氨基酸序列为His - Leu - Leu - Val - Met - X - Gly - Ala - Pro - Glu，表明SITS修饰了钠钾 - ATP酶α亚基上的Lys - 501（X）。