Screening Of A Novel D-arabitol-producing Strain, Optimization Of D-arabitol Production Conditions And Clone And Expression Of Related Key Enzyme Gene

D-arabitol, a five carbon polyol, is widely used in medicine, food and chemical industry. In this study, a new osmophilic yeast identified as Candida sp. H2 by 26S rDNA D1/D2 domain sequences analysis was isolated for D-arabitol production. A weight yield 35% of D-arabitol was achieved in the optimized fermentation medium containing 250 g/L glucose and 10 g/L yeast extract at initial pH 6.0,35℃of culture temperature,200 rpm of agitation,200 mL fermentation medium in a 1000-mL flask of broth content and 1%(v/v) of inoculum size after 96 h fermentation. In addition, a resting Candida sp. H2 cells method for D-arabitol production was investigated and higher weight yield 42% of D-arabitol from glucose was achieved in the conditions consisting of cells with 24 culture time,200 g/L glucose, initial pH 6.0,35℃of culture temperature,200 rpm of agitation,25 mL broth content in a 250-mL flask. Further more, a new D-arabitol 2-dehydrogenase (2-ArDH) gene from Candida sp. H2 which contained 849 nucleotides encoding 282 amino acids with a molecular mass about 30.5 kDa was cloned and sequenced. The amino acid sequence was homogenous to the 2-ArDHs reported which were members of the SDR family. The gene was successfully overexpressed in Escherichia coli BL21(DE3) using a pET-30a vector and the fusion protein was purified with Ni+-NTA column to assay the characteristics of 2-ArDH. The maximum activity of 119.01±1.01 IU/mg for D-arabitol oxidation was obtained at 25℃, pH 9.5. Moreover, this study was first report that E. coli transformed with the gene of 2-ArDH was capable to yield D-arabitol from glucose, displaying the potential of E. coli for synthesizing D-arabitol.