Membrane protein crystallization in meso: lipid type-tailoring of the cubic phase.
Hydrated monoolein forms the cubic-Pn3m mesophase that has been used for in meso crystallization of membrane proteins. The crystals have subsequently provided high-resolution structures by crystallographic means. It is possible that the hosting cubic phase created by monoolein alone, which itself is not a common membrane component, will limit the range of membrane proteins crystallizable by the in meso method. With a view to expanding the range of applicability of the method, we investigated by x-ray diffraction the degree to which the reference cubic-Pn3m phase formed by hydrated monoolein could be modified by other lipid types. These included phosphatidylcholine (PC), phosphatidylethanolamine, phosphatidylserine, cardiolipin, lyso-PC, a polyethylene glycol-lipid, 2-monoolein, oleamide, and cholesterol. The results show that all nine lipids were accommodated in the cubic phase to some extent without altering phase identity. The positional isomer, 2-monoolein, was tolerated to the highest level. The least well tolerated were the anionic lipids, followed by lyso-PC. The others were accommodated to the extent of 20-25 mol %. Beyond a certain concentration limit, the lipid additives either triggered one or a series of phase transitions or saturated the phase and separated out as crystals, as seen with oleamide and cholesterol. The series of phases observed and their order of appearance were consistent with expectations in terms of interfacial curvature changes. The changes in phase type and microstructure have been rationalized on the basis of lipid molecular shape, interfacial curvature, and chain packing energy. The data should prove useful in the rational design of cubic phase crystallization matrices with different lipid profiles that match the needs of a greater range of membrane proteins.
Cherezov
Vadim
V
Chemistry, The Ohio State University, 100 West 18th Avenue, Columbus, OH 43210, USA.
Clogston
Jeffrey
J
Misquitta
Yohann
Y
Abdel-Gawad
Wissam
W
Caffrey
Martin
M
D002308:Cardiolipins / Q000737:chemistry; D002784:Cholesterol / Q000737:chemistry; D003460:Crystallization / Q000379:methods*; D005989:Glycerides / Q000737:chemistry*; D008055:Lipids / Q000737:chemistry*; D008244:Lysophosphatidylcholines / Q000737:chemistry; D008565:Membrane Proteins / Q000138:chemical synthesis / Q000737:chemistry*; D009829:Oleic Acids / Q000737:chemistry; D010713:Phosphatidylcholines / Q000737:chemistry; D010714:Phosphatidylethanolamines / Q000737:chemistry; D010718:Phosphatidylserines / Q000737:chemistry; D013696:Temperature; D014867:Water / Q000737:chemistry; D014961:X-Ray Diffraction / Q000379:methods*
D002308:Cardiolipins; D005989:Glycerides; D008055:Lipids; D008244:Lysophosphatidylcholines; D008565:Membrane Proteins; D009829:Oleic Acids; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010718:Phosphatidylserines; D014867:Water; C006065:We 201; C029407:oleylamide; D002784:Cholesterol; C005953:monoolein