Portal de Periódicos da CAPES

GCN4 Enhances the Stability of the Pore Domain of Potassium Channel KcsA

2009; Elsevier BV; Volume: 96; Issue: 3 Linguagem: Inglês

10.1016/j.bpj.2008.12.1660

1542-0086

Zhiguang Yuchi, Victor P.T. Pau, Daniel S.C. Yang,

Analytical Chemistry and Sensors

KcsA, a prokaryotic potassium channel, is the first channel that has a known crystal structure of the transmembrane domain. The crystal structure of its soluble C-terminal domain, however, still remains elusive. Biophysical and electrophysiological studies have previously implicated the essential roles of the C-terminal domain in pH sensing and in vivo channel assembly. We examined this functional assignment by replacing the C-terminal domain with an artificial tetramerization domain, GCN4-LI. The expression of KcsA is completely abolished when its C-terminal domain is deleted, but it can be rescued by fusion with GCN4-LI. The secondary and quaternary structures of the hybrid channel are very similar to those of the wild type channel according to circular dichroism and gel-filtration analyses. The thermostability of the hybrid channel at pH 8 is similar to that of the wild type but is insensitive to pH changes. This supports the notion that the pH sensor of KcsA is located in the C-terminal domain. Our result is in agreement with the proposed functions of the C-terminal domain and we show that the channel assembly role of the C-terminal domain can be substituted with a non-native tetrameric motif. Since tetramerization domains are found in different families of potassium channels and their presence often enhances the expression of channels, replacement of the elusive C-terminal domains with a known tetrameric scaffold would potentially assist the expression of other potassium channels.