The COOH termini of NBC3 and the 56-kDa H+-ATPase subunit are PDZ motifs involved in their interaction.
The COOH termini of NBC3 and the 56-kDa H+-ATPase subunit are PDZ motifs involved in their interaction.
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DOI：
10.1152/ajpcell.00225.2002
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发表时间：
2003-03-01
影响因子：
5.5
通讯作者：
Kurtz, I
中科院分区：
文献类型：
--
作者：
Pushkin, A;Abuladze, N;Kurtz, I
电中性碳酸氢钠协同转运蛋白3（NBC 3）与空泡H+-ATP酶从肾裂解物中共免疫沉淀。在肾A型和B型闰细胞中，NBC 3与空泡H+-ATP酶共定位。NBC 3的COOH末端和质子泵的56-kDa亚基在这些蛋白质的相互作用的参与进行了研究。合成了完整的和修饰的NBC 3的COOH末端和质子泵的56-kDa亚基，将其偶联到琼脂糖凝胶珠上，并用于拉下肾膜蛋白。质子泵的56-和70-kDa亚基以及含有蛋白质Na+/H+交换调节因子1（NHERF-1）的PDZ结构域都结合到完整的18个氨基酸NBC 3 COOH末端。由5个COOH末端氨基酸截短的肽不结合这些蛋白质。用甘氨酸取代COOH末端亮氨酸阻断了两个质子泵亚基的结合，但不影响NHERF-1的结合。质子泵56 kDa亚基的18个氨基酸COOH末端结合NHERF-1和NBC 3，但截短和修饰的肽不结合。在大鼠肾脏中鉴定了NBC 3（质子泵的56-kDa亚基）和NHERF-1的复合物。这些数据表明，NBC 3的COOH末端和液泡质子泵的56 kDa亚基是PDZ相互作用的基序，这些蛋白质的相互作用是必要的。NHERF-1参与NBC 3和液泡质子泵的相互作用。
The electroneutral sodium bicarbonate cotransporter 3 (NBC3) coimmunoprecipitates from renal lysates with the vacuolar H+-ATPase. In renal type A and B intercalated cells, NBC3 colocalizes with the vacuolar H+-ATPase. The involvement of the COOH termini of NBC3 and the 56-kDa subunit of the proton pump in the interaction of these proteins was investigated. The intact and modified COOH termini of NBC3 and the 56-kDa subunit of the proton pump were synthesized, coupled to Sepharose beads, and used to pull down kidney membrane proteins. Both the 56- and the 70-kDa subunits of the proton pump, as well as a PDZ domain containing protein Na+/H+ exchanger regulatory factor 1 (NHERF-1), were bound to the intact 18 amino acid NBC3 COOH terminus. A peptide truncated by five COOH-terminal amino acids did not bind these proteins. Replacement of the COOH-terminal leucine with glycine blocked binding of both the proton pump subunits but did not affect binding of NHERF-1. The 18 amino acid COOH terminus of the 56-kDa subunit of the proton pump bound NHERF-1 and NBC3, but the truncated and modified peptide did not. A complex of NBC3, the 56-kDa subunit of the proton pump, and NHERF-1 was identified in rat kidney. The data indicate that the COOH termini of NBC3 and the 56-kDa subunit of the vacuolar proton pump are PDZ-interacting motifs that are necessary for the interaction of these proteins. NHERF-1 is involved in the interaction of NBC3 and the vacuolar proton pump.