Manipulating phospholipids for crystallization of a membrane transport protein.
Crystallization is a major bottleneck to obtaining x-ray structures of membrane proteins. By applying an established crystallization protocol for the lactose permease (LacY) of Escherichia coli, a systematic study of the effect of phospholipids (PL) on crystallization of LacY was undertaken. We observe three different crystal forms that diffract to increasingly better resolution in a manner that correlates with the concentration of copurified PL. Consistently, progressive addition of E. coli PL to delipidated LacY leads to different crystal forms. Tetragonal crystals are obtained with improved diffraction quality for a stable mutant by carefully adjusting PL content. Furthermore, crystals of good quality from wild-type LacY, a particularly difficult protein, were also obtained by using same approach. Thus, it is likely that manipulation of PL is a good strategy for predominantly hydrophobic membrane proteins like LacY.
Guan
Lan
L
Department of Physiology, University of California, Los Angeles, CA 90095-1662, USA.
Smirnova
Irina N
IN
Verner
Gill
G
Nagamori
Shushi
S
Nagamoni
Shushi
S
Kaback
H Ronald
HR
D002850:Chromatography, Gel; D003460:Crystallization / Q000379:methods*; D019281:Dimerization; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009004:Monosaccharide Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D010743:Phospholipids / Q000737:chemistry*; D020836:Protein Structure, Quaternary; D027981:Symporters / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D029968:Escherichia coli Proteins; C050221:LacY protein, E coli; D009004:Monosaccharide Transport Proteins; D010743:Phospholipids; D027981:Symporters