A Model Membrane Protein for Binding Volatile Anesthetics
Shixin Ye 1, Joseph Strzalka 1, Inna Y. Churbanova 1, Songyan Zheng 1, Jonas S Johansson 2 and J. Kent Blasie 3*
1 University of Pennsylvania
2 University of Pennsyvania
3 Univ. of Pennsylvania
* To whom correspondence should be addressed. E-mail: jkblasie{at}sas.upenn.edu.
Submitted on August 6, 2004
Revised on August 28, 2004
Accepted on 23 September 2004
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Abstract |
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Earlier work demonstrated that a water-soluble 4-helix bundle protein designed with a cavity in its non-polar core is capable of binding the volatile anesthetic halothane with near-physiological affinity (0.7mM Kd). In order to create a more relevant, model membrane protein receptor for studying the physico-chemical specificity of anesthetic binding, we have synthesized a new protein that builds on the anesthetic-binding, hydrophilic 4-helix bundle and incorporates a hydrophobic domain capable of ion-channel activity, resulting in an amphiphilic 4-helix bundle that forms stable monolayers at the air/water interface. The affinity of the cavity within the core of the bundle for volatile anesthetic binding is decreased by a factor of 4 to 3.1mM Kd as compared to its water-soluble counterpart. Nevertheless, the absence of the cavity within the otherwise identical amphiphilic peptide significantly decreases its affinity for halothane similar to its water-soluble counterpart. Specular X-ray reflectivity shows that the amphiphilic protein orients vectorially in Langmuir monolayers at higher surface pressure with its long axis perpendicular to the interface and possesses a length consistent with its design. This provides a successful starting template for probing the nature of the anesthetic-peptide interaction, as well as a potential model system in structure/function correlation for understanding the anesthetic binding mechanism.
Key Words:
amphiphilic protein, langmuir monolayers, membrane protein, peptide design, volatile anesthetics, x-ray reflectivity
