Dynamic Factors of Bacteriorhodopsin: Regulation Key Points in Helices B, C and F

Abstract

The main objective of this work is the analysis of the structural and functional role of some amino acids in Bacteriorhodopsin.<br/>- Characterization of the motif Thr90-Pro91 in the centre of Helix C of Bacteriorhodopsin. Because Thr90 interacts with Asp115 through a hydrogen bond and with the retinal through hydrophobic interaction, the mutants T90V and D115A were taken into account. The following objectives were settled:<br/>Design and construct the Bacteriorhodopsin mutants P91A, T90A,T90V and D115A by site directed mutagenesis.<br/>Express the mutants in the purple membrane of Halobacterium salinarum.<br/>Analyze the role of Thr90 and Pro91 in the properties of the protein and in the proton pumping function in particular through the study of the mutants.<br/>Infer a role to the motif Thr90-Pro91 in Bacteriorhodopsin.<br/>- Analyze the role of the other two Prolines located in transmembrane helices in BR, and to extend this role to other Transmembrane Proteins. The subsequent objectives were proposed:<br/>Design and construct the Bacteriorhodopsin mutants P50A and P186A by site directed mutagenesis.<br/>Express the mutants in the purple membrane of Halobacterium salinarum.<br/>Characterize the role of Pro50, and Pro186 in Bacteriorhodopsin properties and proton pumping through the analysis of the mutants.<br/>Infer a role to Helix-embedded prolines of Bacteriorhodopsin in particular and to transmembrane helices of proteins in general.