Overexpression, purification, crystallization and preliminary X-ray analysis of putative molybdenum cofactor biosynthesis protein C (MoaC2) from Mycobacterium tuberculosis H37Rv.
Rv0864 (MoaC2) from Mycobacterium tuberculosis is one of the enzymes in the molybdenum cofactor (Moco) biosynthesis pathway. Together with MoaA, MoaC is involved in the conversion of guanosine triphosphate (GTP) to precursor Z, the first step in Moco synthesis. Full-length MoaC2 (17.5 kDa, 167 residues) was cloned in Escherichia coli and purified to homogeneity. Crystals of recombinant M. tuberculosis MoaC2 were grown by vapour diffusion using a hanging-drop setup. Diffracting crystals grew in a condition in which 3 µl protein solution at 10.5 mg ml(-1) was mixed with 1.5 µl reservoir solution (0.025 M potassium sodium tartrate tetrahydrate pH 8.0) and equilibrated against 1000 µl reservoir solution. Diffraction data extending to 2.5 Å resolution were collected at 100 K. The crystal belonged to the cubic space group P2(1)3, with unit-cell parameter 94.5 Å. Matthews coefficient (V(M)) calculations suggested the presence of two molecules in the asymmetric unit, corresponding to a solvent content of about 39%. Molecular-replacement calculations using the E. coli homologue as the search model gave an unambiguous solution.
Molecular and Structural Biology Division, CSIR - Central Drug Research Institute, Chattar Manzil Palace, Lucknow 226 001, India.
Shubhra
S
Srivastava
Vijay Kumar
VK
Srivastava
Ashish
A
Arora
J Venkatesh
JV
Pratap
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification; D003460:Crystallization; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000737:chemistry*; D009687:Nuclear Proteins / Q000737:chemistry* / Q000302:isolation & purification; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment
D001426:Bacterial Proteins; D009687:Nuclear Proteins