The molecular forces that stabilize membrane protein structure are poorly understood. To investigate these forces we introduced alanine substitutions at 24 positions in the B helix of bacteriorhodopsin and examined their effects on structure and stability. Although most of the results can be rationalized in terms of the folded structure, there are a number of surprises. (1) We find a remarkably high frequency of stabilizing mutations (17%), indicating that membrane proteins are not highly optimized for stability. (2) Helix B is kinked, with the kink centered around Pro50. The P50A mutation has no effect on stability, however, and a crystal structure reveals that the helix remains bent, indicating that tertiary contacts dominate in the distortion of this helix. (3) We find that the protein is stabilized by about 1kcal/mol for every 38A(2) of surface area buried, which is quite similar to soluble proteins in spite of their dramatically different environments. (4) We find little energetic difference, on average, in the burial of apolar surface or polar surface area, implying that van der Waals packing is the dominant force that drives membrane protein folding. Study holds ProTherm entries: 17314, 17315, 17316, 17317, 17318, 17319, 17320, 17321, 17322, 17323, 17324, 17325, 17326, 17327, 17328, 17329, 17330, 17331, 17332, 17333, 17334, 17335, 17336, 17337 Extra Details: 15 mM DMPC and 16 mM CHAPSO were added in the experiment. bacteriorhodopsin; hydrogen bond; van der Waals; protein folding; helix kink
Submitter: Connie Wang
Submission Date: April 24, 2018, 8:49 p.m.
|Number of data points||24|
|Proteins||Bacteriorhodopsin ; Bacteriorhodopsin|
|Assays/Quantities/Protocols||Experimental Assay: ddG_H2O|
|Libraries||Mutations for sequence EAQITGRPEWIWLALGTALMGLGTLYFLVKGMGVSDPDAKKFYAITTLVPAIAFTMYLSMLLGYGLTMVPFGGEQNPIYWARYADWLFTTPLLLLDLALLVDADQGTILALVGADGIMIGTGLVGALTKVYSYRFVWWAISTAAMLYILYVLFFGFTSKAESMRPEVASTFKVLRNVTVVLWSAYPVVWLIGSEGAGIVPLNIETLLFMVLDVSAKVGFGLILLRSRAIFGEAEAPEPSAGDGAAATS|