Conversion of a porin-like peptide channel into a gramicidin-like channel by glycine to D-alanine substitutions

Abstract

The β-barrel and β-helix formation, as in porins and gramicidin, respectively, represent two distinct mechanisms for ion channel formation by β-sheet proteins in membranes. The design of β-barrel proteins is difficult due to incomplete understanding of the basic principles of folding. The design of gramicidin-like β-helix relies on an alternating pattern of L- and D-amino acid sequences. Recently we noticed that a short β-sheet peptide (xSxG)6, can form porin-like channels via self-association in membranes. Here, we proposed that glycine to D-alanine substitutions of the N-formyl-(xSxG)6 would transform the porin-like channel into a gramicidin-like β12-helical channel. The requirement of an N-formyl group for channel activity, impermeability to cations with a diameter >4 Å, high monovalent cation selectivity, and the absence of either voltage gating or subconductance states upon D-alanine substitution support the idea of a gramicidin-like channel. Moreover, the circular dichroism spectrum in membranes is different, indicating a change in regular β-sheet backbone structure. The conversion of a complex porin-like channel into a gramicidin-like channel provides a link between two different mechanisms of β-sheet channel formation in membranes and emphasizes the importance of glycine and D-amino acid residues in protein folding and function and in the engineering of ion channels. © 2006 by the Biophysical Society.