Effects of glycine substitutions on the structure and function of gramicidin a channels.
Tryptophan residues often are found at the lipid-aqueous interface region of membrane-spanning proteins, including ion channels, where they are thought to be important determinants of protein structure and function. To better understand how Trp residues modulate the function of membrane-spanning channels, we have examined the effects of Trp replacements on the structure and function of gramicidin A channels. Analogues of gramicidin A in which the Trp residues at positions 9, 11, 13, and 15 were sequentially replaced with Gly were synthesized, and the three-dimensional structure of each analogue was determined using a combination of two-dimensional NMR techniques and distance geometry-simulated annealing structure calculations. Though Trp --> Gly substitutions destabilize the beta6.3-helical gA channel structure, it is possible to determine the structure of analogues with Trp --> Gly substitutions at positions 11, 13, and 15, but not for the analogue with the Trp --> Gly substitution at position 9. The Gly11-, Gly13-, and Gly15-gA analogues form channels that adopt a backbone fold identical to that of native gramicidin A, with only small changes in the side chain conformations of the unsubstituted residues. Single-channel current measurements show that the channel function and lifetime of the analogues are significantly affected by the Trp --> Gly replacements. The conductance variations appear to be caused by sequential removal of the Trp dipoles, which alter the ion-dipole interactions that modulate ion movement. The lifetime variations did not appear to follow a clear pattern.
Jordan
J B
JB
Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, Arkansas 72701, USA.
Shobana
S
S
Andersen
O S
OS
Hinton
J F
JF
D019943:Amino Acid Substitution; D005998:Glycine / Q000737:chemistry*; D006096:Gramicidin / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular
D006096:Gramicidin; D005998:Glycine