Open AccessOverexpression, purification, crystallization and preliminary crystallographic studies of a hyperthermophilic adenylosuccinate synthetase from Pyrococcus horikoshii OT3
Author(s) -
Wang Xiaoying,
Akasaka Ryogo,
Takemoto Chie,
Morita Satoshi,
Yamaguchi Machiko,
Terada Takaho,
Shirozu Mikako,
Yokoyama Shigeyuki,
Chen Shilin,
Si Shuyi,
Xie Yong
Publication year - 2011
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
ISSN - 1744-3091
DOI - 10.1107/s174430911104108x
Subject(s) - pyrococcus horikoshii , thermophile , archaea , dimer , inosine monophosphate , crystallography , crystallization , orthorhombic crystal system , biochemistry , chemistry , biology , stereochemistry , nucleotide , enzyme , crystal structure , gene , organic chemistry
Adenylosuccinate synthetase (AdSS) is a ubiquitous enzyme that catalyzes the first committed step in the conversion of inosine monophosphate (IMP) to adenosine monophosphate (AMP) in the purine‐biosynthetic pathway. Although AdSS from the vast majority of organisms is 430–457 amino acids in length, AdSS sequences isolated from thermophilic archaea are 90–120 amino acids shorter. In this study, crystallographic studies of a short AdSS sequence from Pyrococcus horikoshii OT3 (PhAdSS) were performed in order to reveal the unusual structure of AdSS from thermophilic archaea. Crystals of PhAdSS were obtained by the microbatch‐under‐oil method and X‐ray diffraction data were collected to 2.50 Å resolution. The crystal belonged to the trigonal space group P 3 2 12, with unit‐cell parameters a = b = 57.2, c = 107.9 Å. There was one molecule per asymmetric unit, giving a Matthews coefficient of 2.17 Å 3 Da −1 and an approximate solvent content of 43%. In contrast, the results of native polyacrylamide gel electrophoresis and analytical ultracentrifugation showed that the recombinant PhAdSS formed a dimer in solution.