Towards a three-alpha-helix bundle protein that binds volatile general anesthetics.
The general anesthetics halothane and chloroform are capable of binding to synthetic water-soluble four-alpha-helix bundles, which model the putative in vivo receptors. In this study, we investigate the binding of these anesthetics to synthetic water-soluble three-alpha-helix bundles. A series of variants containing up to four X-to-Ala and up to four X-to-Met substitutions was made; and the effect of these substitutions on structure, stability and anesthetic binding affinity was examined. Generally, the amount of alpha-helix and the stability of the three-alpha-helix bundles decreased as the number of X-to-Ala substitutions increased. A concomitant red-shift in tryptophan fluorescence lambdamax was seen, suggesting an increased flexibility of the native structure. Up to four X-to-Met substitutions had little effect on the amount of alpha-helix, but an increase in tryptophan lambdamax was seen for the variants with three and four methionine substitutions. The exceptions were a) a variant with a clustering of alanine and methionine residues at one end of the three-alpha-helix bundle, suggesting a gate structure that can admit ligand molecules; and b) a variant with a single Leu35Ala substitution, suggesting that at select positions, the size of the side chain is important for defining anesthetic binding affinity.
Manderson
Gavin A
GA
University of Pennsylvania, Department of Anesthesia and the Johnson Research Foundation, Philadelphia, PA 19104, USA.
Johansson
Jonas S
JS
D000595:Amino Acid Sequence; D018681:Anesthetics, General / Q000378:metabolism*; D002352:Carrier Proteins / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D002725:Chloroform / Q000378:metabolism; D002942:Circular Dichroism; D006221:Halothane / Q000378:metabolism; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary
D018681:Anesthetics, General; D002352:Carrier Proteins; D008565:Membrane Proteins; D002725:Chloroform; D006221:Halothane