Korean Journal of Microbiology (미생물학회지)

The Microbiological Society of Korea (한국미생물학회)
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Enzymatic Characterization of Salmonella typhimurium Mannitol Dehydrogenase Expressed in Escherichia coli

Salmonella typhimurium에서 유래한 Mannitol Dehydrogenase 유전자의 대장균 내 발현 및 효소특성 규명

  • Jang, Myoung-Uoon (Department of Food Science and Technology, Chungbuk National University) ;
  • Park, Jung-Mi (Department of Food Science and Technology, Chungbuk National University) ;
  • Kim, Min-Jeong (Department of Food Science and Technology, Chungbuk National University) ;
  • Kang, Jung-Hyun (Department of Food Science and Technology, Chungbuk National University) ;
  • Lee, So-Won (Department of Food Science and Technology, Chungbuk National University) ;
  • Kim, Tae-Jip (Department of Food Science and Technology, Chungbuk National University)
  • Received : 2012.06.05
  • Accepted : 2012.06.25
  • Published : 2012.06.30
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Abstract

A mannitol dehydrogenase (StMDH) gene was cloned from Salmonella typhimurium LT2 (KCTC 2421) and overexpressed in Escherichia coli. It has a 1,467 bp open reading frame encoding 488 amino acids with deduced molecular mass of 54 kDa, which shares approximately 36% of amino acid identity with known long-chain dehydrogenase/reductatse (LDR) family enzymes. The recombinant StMDH showed the highest activity at $30^{\circ}C$, and pH 5.0 and 10.0 for D-fructose reduction and D-mannitol oxidation, respectively. On the contrary, it has no activity on glucose, galactose, xylose, and arabinose. StMDH can catalyze the oxidative/reductive reactions between D-fructose and D-mannitol only in the presence of $NAD^+$/NADH as coenzymes. These results indicate that StMDH is a typical $NAD^+$/NADH-dependent mannitol dehydrogenase (E.C. 1.1.1.67).

Salmonella typhimurium LT2 (KCTC 2421)로부터 mannitol dehydrogenase (StMDH)로 추정되는 유전자를 클로닝하고, 대장균에서 대량 발현하였다. 이 유전자는 488개의 아미노산 서열(약 54 kDa)을 암호화하는 1,467 bp의 염기로 구성되며, 이미 보고된 long-chain dehydrogenase/ reductase (LDR) 계열 효소들과 약 36%의 아미노산 서열 상동성을 나타내었다. 재조합 StMDH의 최적 반응온도는 $30^{\circ}C$이며, pH 5.0의 조건에서 최대의 D-fructose 환원활성, 그리고 pH 10.0에서 최대의 D-mannitol 산화활성을 보인다. 반면에 glucose, galactose, xylose, arabinose 등의 기질에 대해서는 활성을 보이지 않았다. 이 효소는 $NAD^+$/NADH 존재 하에서만 산화 환원 활성을 가지며, $NADP^+$/NADPH는 조효소로 이용하지 못하였다. 결론적으로 StMDH는 전형적인 $NAD^+$/NADH 의존형의 mannitol dehydrogenase (EC 1.1.1.67)임을 확인하였다.

Keywords

References

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