title: Enzymatic characterization of isocitrate dehydrogenase from an emerging zoonotic pathogen Streptococcus suis.
abstract: Streptococcus suis, a Gram-positive coccus, is an emerging zoonotic pathogen for both humans and pigs, but little is known about the properties of its metabolic enzymes. Isocitrate dehydrogenase (IDH) is a key regulatory enzyme in the citric acid cycle that catalyzes the oxidative decarboxylation of isocitrate yielding α-ketoglutarate and NAD(P)H. Here, we report the overexpression and enzymatic characterization of IDH from S. suis Serotype 2 Chinese highly virulent strain 05ZYH33 (SsIDH). The molecular weight of SsIDH was estimated to be 74 kDa by gel filtration chromatography, suggesting a homodimeric structure. Additionally, SsIDH was divalent cation-dependent and Mg(2+) was found to be the most effective cation. The optimal pH of SsIDH was 7.0 (Mn(2+)) and 8.5 (Mg(2+)), and the maximum activity was around 30 °C (Mn(2+)) and 50 °C (Mg(2+)), respectively. Heat inactivation studies showed that SsIDH retained 50% activity after 20 min of incubation at 49 °C. Sequence comparison revealed that SsIDH had a significantly homologous identity to bacterial homodimeric IDHs. The recombinant SsIDH displayed a 117-fold (k(cat)/K(m)) preference for NAD(+) over NADP(+) with Mg(2+), and a 80-fold greater specificity for NAD(+) than NADP(+) with Mn(2+). Therefore, SsIDH has remarkably high coenzyme preference toward NAD(+). This current work is expected to shed light on the functions of metabolic enzymes in S. suis and provide useful information for SsIDH to be considered as a possible candidate for serological diagnostics and detection of S. suis infection.
authors.affiliation: Key Laboratory of Molecular Evolution and Biodiversity, and Institute of Molecular Biology and Biotechnology, Anhui Normal University, Wuhu 241000, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Wang
authors.forename: Mingming
authors.initials: M
authors.lastname: Jin
authors.forename: Ruirui
authors.initials: R
authors.lastname: Su
authors.forename: Ping
authors.initials: P
authors.lastname: Song
authors.forename: Ming
authors.initials: M
authors.lastname: Wang
authors.forename: Guoping
authors.initials: G
authors.lastname: Zhu
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D007521:Isocitrate Dehydrogenase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D017011:Streptococcus suis / Q000201:enzymology* / Q000472:pathogenicity
chemical_list: D001426:Bacterial Proteins; D009243:NAD; D009249:NADP; D007521:Isocitrate Dehydrogenase