Browsing by Author "Rani, Priya"

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    Cloning, expression, purification and crystallization of a novel GlcNAc metabolic protein, gig2 (duf1479) from pathogenic fungus Candida albicans
    (Proteomics Society, India (PSI), 2017) Rani, Priya; Gautam, Gunjan; Rao, Kongara Hanumantha; Ghosh, Swagata; Gourinath, Samudrala; Dhar, Suman Kumar; Datta, Asis
    N-acetylglucosamine (GlcNAc), an alternative sugar, is emerging as an important molecule having a multifarious role in Candida albicans including a major role in signaling. GlcNAc Inducible Gene 2, GIG2 is one of the highly upregulated genes in GlcNAc grown cells in C. albicans. Our earlier studies show the involvement of Gig2 in the formation of N-acetylneuraminic (NANA) acid from GlcNAc-6-phosphate through an understudied route. The crystal structure of Gig2 would help us in determining the exact reaction that this enzyme catalyzes. Here the cloning, expression, purification and crystallization of this protein are reported along with preliminary X-ray crystallographic analysis at 2.4Å resolution. The crystal belonged to P2 1 space group, with unit cell parameters a=59.59, b= 54.43, c= 73.29Å; α = 90°, β = 102.7° and γ = 90°. The structure was solved using PDB ID 2CSG as a template which has only 27% identity. Molecular replacement yielded a solution with LLG score of 87. The structure is currently under further refinement.
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    Crystal structure of Gig2 protein from Candida albicans provides a structural insight into DUF1479 family oxygenases
    (Elsevier B.V., 2020) Rani, Priya; Gautam, Gunjan; Anwar, Tamanna; Gourinath, Samudrala; Datta, Asis
    Candida albicans, GlcNAc inducible gene 2 (Gig2) is important for its virulence, oxidative stress adaptation and is supposed to be a part of the undefined GlcNAc metabolic network. On the basis of sequence homology, Gig2 is classified as a putative oxidoreductase of DUF1479 family (Domain of Unknown Function) with no reported structure and function. In this work, we have elucidated the crystal structure of Gig2 protein using X-ray crystallography at 1.7 Å resolution. Crystals were improved using successive macro-seeding technique. Structure was solved by molecular replacement using a template (PDBID: 2CSG) with a mere 27% identity with Gig2, followed by manual and automated model building. Gig2 exists as a monomer with a single large DUF1479 domain and is composed of a cupin like double stranded β-helix (DBSH) core fold with Iron (Fe) in the active site. This is the first report of DUF1479 family proteins which identifies and highlights its unique structural features. Crystal structure elucidation, structural comparisons, and docking studies proposes Gig2 as a non-heme Fe (II) containing 2-oxoglutarate dependent oxygenase.