4MSW

Y78 ester mutant of KcsA in high K+

Summary for 4MSW

• Entry DOI: 10.2210/pdb4msw/pdb
• Descriptor: ANTIBODY FAB FRAGMENT HEAVY CHAIN, Monoclonal 11D8 anti-human butyrylcholinesterase (BChE) light chain, pH-gated potassium channel KcsA, ... (6 entities in total)
• Functional Keywords: membrane protein, channel, ester, unnatural amino acid, transport protein
• Biological source: Mus musculus; More
• Cellular location: Cell membrane; Multi-pass membrane protein: 4MSW
• Total number of polymer chains: 3
• Total formula weight: 58696.32

Authors

Matulef, K.,Valiyaveetil, F.I. (deposition date: 2013-09-18, release date: 2013-10-30, Last modification date: 2025-03-26)

Primary citation

Matulef, K.,Komarov, A.G.,Costantino, C.A.,Valiyaveetil, F.I.
Using protein backbone mutagenesis to dissect the link between ion occupancy and C-type inactivation in K+ channels.
Proc.Natl.Acad.Sci.USA, 110:17886-17891, 2013

PubMed Abstract: K(+) channels distinguish K(+) from Na(+) in the selectivity filter, which consists of four ion-binding sites (S1-S4, extracellular to intracellular) that are built mainly using the carbonyl oxygens from the protein backbone. In addition to ionic discrimination, the selectivity filter regulates the flow of ions across the membrane in a gating process referred to as C-type inactivation. A characteristic of C-type inactivation is a dependence on the permeant ion, but the mechanism by which permeant ions modulate C-type inactivation is not known. To investigate, we used amide-to-ester substitutions in the protein backbone of the selectivity filter to alter ion binding at specific sites and determined the effects on inactivation. The amide-to-ester substitutions in the protein backbone were introduced using protein semisynthesis or in vivo nonsense suppression approaches. We show that an ester substitution at the S1 site in the KcsA channel does not affect inactivation whereas ester substitutions at the S2 and S3 sites dramatically reduce inactivation. We determined the structure of the KcsA S2 ester mutant and found that the ester substitution eliminates K(+) binding at the S2 site. We also show that an ester substitution at the S2 site in the KvAP channel has a similar effect of slowing inactivation. Our results link C-type inactivation to ion occupancy at the S2 site. Furthermore, they suggest that the differences in inactivation of K(+) channels in K(+) compared with Rb(+) are due to different ion occupancies at the S2 site.

PubMed: 24128761
DOI: 10.1073/pnas.1314356110

Experimental method

X-RAY DIFFRACTION (2.06 Å)