Role of Tryptophan Residues in Gramicidin Channel Organization and Function

¹ Centre for Cellular & Molecular Biology
² Department of Chemistry and Biochemistry, University of Arkansas

^* To whom correspondence should be addressed. E-mail: amit{at}ccmb.res.in.

Submitted on October 22, 2007
Revised on November 26, 2007
Accepted on 28 February 2008

	Abstract

The linear peptide gramicidin forms prototypical ion channels specific for monovalent cations and has been used extensively to study the organization, dynamics and function of membrane-spanning channels. The tryptophan residues in gramicidin channels are crucial for maintaining the structure and function of the channel. We have explored the structural basis for the reduction in channel conductance in the case of single tryptophan analogues of gramicidin with three Trp hydrophobic substitutions using a combination of fluorescence approaches which include red edge excitation shift (REES) and membrane penetration depth analysis, size-exclusion chromatography and CD spectroscopy. We show here that the gramicidin analogues containing single tryptophan residues adopt a mixture of non-channel and channel conformations, as evident from analysis of membrane penetration depth, size-exclusion chromatography and backbone CD data. The results correlate well with the reduction in single channel conductance and channel-forming propensity exhibited by similar single-Trp analogues. These results could be potentially useful in analyzing the effect of tryptophan substitution on the functioning of other ion channels and membrane proteins.

Key Words: Gramicidin, REES, ion channel, membrane penetration depth, size exclusion chromatography, tryptophan