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Cloning, expression, purification, crystallization and preliminary X-ray crystallographic study of the putative SAICAR synthetase (PH0239) from Pyrococcus horikoshii OT3

Manjunath, K and Jeyakanthan, J and Nakagawa, N and Shinkai, A and Yoshimura, M and Kuramitsu, S and Yokoyama, S and Sekar, K (2010) Cloning, expression, purification, crystallization and preliminary X-ray crystallographic study of the putative SAICAR synthetase (PH0239) from Pyrococcus horikoshii OT3. In: Acta Crystallographica Section F, 66 (Part 2). pp. 180-183.

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Abstract

The study of proteins involved in de novo biosynthesis of purine nucleotides is central in the development of antibiotics and anticancer drugs. In view of this, a protein from the hyperthermophile Pyrococcus horikoshii OT3 was isolated, purified and crystallized using the microbatch method. Its primary structure was found to be similar to that of SAICAR synthetase, which catalyses the seventh step of de novo purine biosynthesis. A diffraction-quality crystal was obtained using Hampton Research Crystal Screen II condition No. 34, consisting of 0.05 M cadmium sulfate hydrate, 0.1 M HEPES buffer pH 7.5 and 1.0 M sodium acetate trihydrate, with 40%(v/v) 1,4-butanediol as an additive. The crystal belonged to space group P3(1), with unit-cell parameters a = b = 95.62, c = 149.13 angstrom. Assuming the presence of a hexamer in the asymmetric unit resulted in a Matthews coefficient (V-M) of 2.3 angstrom(3) Da(-1), corresponding to a solvent content of about 46%. A detailed study of this protein will yield insights into structural stability at high temperatures and should be highly relevant to the development of antibiotics and anticancer drugs targeting the biosynthesis of purine nucleotides.

Item Type: Journal Article
Additional Information: Copyright of this article belongs to John Wiley and Sons.
Keywords: SAICAR synthetase; PH0239; Pyrococcus horikoshii OT3; purine biosynthesis.
Department/Centre: Division of Information Sciences > Supercomputer Education & Research Centre
Division of Information Sciences > BioInformatics Centre
Date Deposited: 15 Jul 2010 07:07
Last Modified: 19 Sep 2010 06:10
URI: http://eprints.iisc.ernet.in/id/eprint/28913

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