期刊
JOURNAL OF GENERAL PHYSIOLOGY
卷 126, 期 6, 页码 563-570出版社
ROCKEFELLER UNIV PRESS
DOI: 10.1085/jgp.200509417
关键词
-
类别
资金
- NIGMS NIH HHS [R37 GM031768, GM-31768, R01 GM031768] Funding Source: Medline
CLC-ec1 is a prokaryotic CLC-type Cl-/H+ exchange transporter. Little is known about the mechanism of H+ coupling to Cl-. A critical glutamate residue, E148, was previously shown to be required for Cl-/H+ exchange by mediating proton transfer between the protein and the extracellular solution. To test whether an analogous H+ acceptor exists near the intracellular side of the protein, we performed a mutagenesis scan of inward-facing carboxyl-bearing residues and identified E203 as the unique residue whose neutralization abolishes H+ coupling to Cl- transport. Glutamate at this position is strictly conserved in all known CLCs of the transporter subclass, while valine is always found here in CLC channels. The x-ray crystal structure of the E203Q mutant is similar to that of the wild-type protein. Cl- transport rate in E203Q is inhibited at neutral pH, and the double mutant, E148A/E203Q, shows maximal Cl- transport, independent of pH, as does the single mutant E148A. The results argue that substrate exchange by CLC-ec1 involves two separate but partially overlapping permeation pathways, one for Cl- and one for H+. These pathways are congruent from the protein's extracellular surface to E148, and they diverge beyond this point toward the intracellular side. This picture demands a transport mechanism fundamentally different from familiar alternating-access schemes.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据