The effects of a protein osmolyte on the stability of the integral membrane protein glycerol facilitator.
Osmolytes are naturally occurring molecules used by a wide variety of organisms to stabilize proteins under extreme conditions of temperature, salinity, hydrostatic pressure, denaturant concentration, and desiccation. The effects of the osmolyte trimethylamine N-oxide (TMAO) as well as the influence of detergent head group and acyl chain length on the stability of the Escherichia coli integral membrane protein glycerol facilitator (GF) tetramer to thermal and chemical denaturation by sodium dodecyl sulphate (SDS) are reported. TMAO promotes the association of the normally tetrameric α-helical protein into higher order oligomers in dodecyl-maltoside (DDM), but not in tetradecyl-maltoside (TDM), lyso-lauroylphosphatidyl choline (LLPC), or lyso-myristoylphosphatidyl choline (LMPC), as determined by dynamic light scattering (DLS); an octameric complex is particularly stable as indicated by SDS polyacrylamide gel electrophoresis. TMAO increases the heat stability of the GF tetramer an average of 10 °C in the 4 detergents and also protects the protein from denaturation by SDS. However, it did not promote re-association to the tetramer when added to SDS-dissociated protein. TMAO also promotes the formation of rod-like detergent micelles, and DLS was found to be useful for monitoring the structure of the protein and the redistribution of detergent during thermal dissociation of the protein. The protein is more thermally stable in detergents with the phosphatidylcholine head group (LLPC and LMPC) than in the maltoside detergents. The implications of the results for osmolyte mechanism, membrane protein stability, and protein-protein interactions are discussed.
Department of Chemistry, University of Manitoba, Winnipeg, MB R3T 2N2, Canada.
Simon
S
Baturin
Jamie J
JJ
Galka
Hadeesha
H
Piyadasa
S
S
Gajjeraman
Joe D
JD
O'Neil
D020346:Aquaporins / Q000737:chemistry* / Q000378:metabolism; D003902:Detergents / Q000737:chemistry*; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D008744:Methylamines / Q000737:chemistry*; D055550:Protein Stability; D020836:Protein Structure, Quaternary
D020346:Aquaporins; D003902:Detergents; D029968:Escherichia coli Proteins; D008744:Methylamines; C078043:GlpF protein, E coli; C005855:trimethyloxamine
SDS; TMAO; aquaporines; aquaporins; detergent; détergent; facilitateur de glycérol; glycerol facilitator; membrane protein; osmolyte; protéine membranaire; stabilité thermique; tetramer; thermal stability; tétramère