title: Cloning, overexpression, purification and preliminary X-ray analysis of a feast/famine regulatory protein (Rv2779c) from Mycobacterium tuberculosis H37Rv.
abstract: Rv2779c from Mycobacterium tuberculosis is a feast/famine regulatory protein. This class of proteins are also known as the leucine-responsive regulatory protein/asparagine synthase C family (Lrp/AsnC) of transcriptional regulators and are known to be involved in various metabolic processes in bacteria and fungi. They contain a RAM (regulator of amino-acid metabolism) domain that is rarely found in humans and acts as the oligomerization domain. Since the oligomeric status is often linked to the particular functional role in these proteins, binding of ligands to the domain can elicit specific functional responses. Full-length Rv2779c corresponding to a molecular mass of 19.8 kDa and 179 residues was cloned and purified to homogeneity following transformation into Escherichia coli C41 (DE3) cells. Crystals were grown by vapour diffusion using the hanging-drop method. Diffraction data extending to 2.8 Å resolution were collected from a single crystal that belonged to space group P2(1)2(1)2, with unit-cell parameters a = 99.6, b = 146.0, c = 49.9 Å. Matthews coefficient (VM) calculations suggest that four molecules are present in the asymmetric unit, corresponding to a solvent content of ∼46%. Molecular-replacement calculations using the crystal structure of a homologue, Rv3291c, as the search model gave an unambiguous solution corresponding to four subunits in the asymmetric unit.
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, B.S/10/1, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow 226 031, India.
authors.forename: Abhishek
authors.initials: A
authors.lastname: Dey
authors.affiliation: Molecular and Structural Biology Division, CSIR-Central Drug Research Institute, B.S/10/1, Sector-10, Jankipuram Extension, Sitapur Road, Lucknow 226 031, India.
authors.forename: Ravishankar
authors.initials: R
authors.lastname: Ramachandran
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004058:Diffusion; D004591:Electrophoresis, Polyacrylamide Gel; D009169:Mycobacterium tuberculosis / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins
keywords: Mycobacterium tuberculosis; Rv2779c; feast/famine regulatory proteins