Cloning and characterization of a galactitol 2-dehydrogenase from Rhizobium legumenosarum and its application in d-tagatose production

Sujit Sadashiv Jagtap, Ranjitha Singh, Yun Chan Kang, Huimin Zhao, Jung Kul Lee

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Galactitol 2-dehydrogenase (GDH) belongs to the protein subfamily of short-chain dehydrogenases/reductases and can be used to produce optically pure building blocks and for the bioconversion of bioactive compounds. An NAD+-dependent GDH from Rhizobium leguminosarum bv. viciae 3841 (RlGDH) was cloned and overexpressed in Escherichia coli. The RlGDH protein was purified as an active soluble form using His-tag affinity chromatography. The molecular mass of the purified enzyme was estimated to be 28kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 114kDa by gel filtration chromatography, suggesting that the enzyme is a homotetramer. The enzyme has an optimal pH and temperature of 9.5 and 35°C, respectively. The purified recombinant RlGDH catalyzed the oxidation of a wide range of substrates, including polyvalent aliphatic alcohols and polyols, to the corresponding ketones and ketoses. Among various polyols, galactitol was the preferred substrate of RlGDH with a Km of 8.8mM, kcat of 835min-1 and a kcat/Km of 94.9min-1mM-1. Although GDHs have been characterized from a few other sources, RlGDH is distinguished from other GDHs by its higher specific activity for galactitol and broad substrate spectrum, making RlGDH a good choice for practical applications.

Original languageEnglish
Pages (from-to)44-51
Number of pages8
JournalEnzyme and Microbial Technology
Volume58-59
DOIs
Publication statusPublished - 2014 May 10

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Keywords

  • Galactitol 2-dehydrogenase
  • Homology modeling
  • SDR enzyme
  • Sugar alcohols

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Biochemistry
  • Applied Microbiology and Biotechnology

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