Expression, purification, crystallization and preliminary X-ray diffraction analysis of an essential lipoprotein implicated in cell-wall biosynthesis in Mycobacteria.
Mycobacterium tuberculosis is a renewed cause of devastation in the developing world. Critical to the success of this re-emerging pathogen is its unusual waxy cell wall, which is rich in rare components including lipoarabinomannan (LAM) and its precursors, the phosphatidylinositol mannosides (PIMs). Balanced synthesis of these related glycolipids is intrinsic to both cell-wall integrity and virulence in M. tuberculosis and presents a promising, albeit poorly defined, therapeutic target. Here, the expression, purification and crystallization of an essential 600-amino-acid lipoprotein, LpqW, implicated in this process are reported. Crystals of LpqW were grown using 20-24%(w/v) PEG 4000, 8-16%(v/v) 2-propanol, 100 mM sodium citrate pH 5.5 and 10 mM DTT. A complete data set was collected at 2.4 A using synchrotron radiation on a crystal belonging to space group C222, with unit-cell parameters a = 188.57, b = 312.04, c = 104.15 A. Structure determination is under way.
Marland
Zara
Z
Department of Biochemistry and Molecular Biology, School of Biomedical Sciences, Monash University, Clayton, Victoria 3800, Australia.
Beddoe
Travis
T
Zaker-Tabrizi
Leyla
L
Coppel
Ross L
RL
Crellin
Paul K
PK
Rossjohn
Jamie
J
D001426:Bacterial Proteins / Q000737:chemistry*; D002473:Cell Wall / Q000378:metabolism; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004591:Electrophoresis, Polyacrylamide Gel; D008070:Lipopolysaccharides / Q000737:chemistry; D008074:Lipoproteins / Q000737:chemistry*; D008362:Mannosides / Q000737:chemistry; D009169:Mycobacterium tuberculosis / Q000378:metabolism*; D010716:Phosphatidylinositols / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry; D017434:Protein Structure, Tertiary; D014961:X-Ray Diffraction
D001426:Bacterial Proteins; D008070:Lipopolysaccharides; D008074:Lipoproteins; C510627:LpqW protein, Mycobacterium tuberculosis; D008362:Mannosides; D010716:Phosphatidylinositols; C050016:lipoarabinomannan; D011092:Polyethylene Glycols