Genetically engineered bacteria and starter for the dehydrogenation of the 1 and 2 positions of the a ring of steroidal compounds

Abstract

The invention relates to a genetically engineered bacterium and a leavening agent for dehydrogenation of steroid A ring 1,2 positions. The host bacterium of the genetically engineered bacterium is Arthrobacter simplex, the preservation number of which is ACCC 12070. The genetically engineered bacterium contains a recombinant gene fragment as shown in Sequence 2. The fermentation substrate of the genetically engineered bacterium contains cortisone acetate, fluoride cortical hormone and 4-AD. The constructed engineered bacterium provided by the invention performs fermentation by constant temperature ventilation stirring on the basis of yeast extract, phosphate, corn steep liquor and glucose containing nutritional ingredients. When the concentration of steroids is not higher than 10%(m / V), one time addition of steroid substrate is adopted, and?biological dehydrogenation conversion rate can reach to more than 95% in?32-48 hours. When the concentration of steroids is higher than 10%, continuous addition of steroid substrate is adopted, and biological dehydrogenation conversion rate can reach to more than 95% within 60 hours.

Description

technical field [0001] The invention belongs to the field of genetic engineering and microbial fermentation, and relates to Arthrobacter simplex used for high-efficiency transformation of steroidal compounds (dehydrogenation of 1 and 2 positions of A ring), especially a method for the dehydrogenation of 1 and 2 positions of A ring of steroid compounds. Hydrogen genetically engineered bacteria and starter. [0002] technical background [0003] Steroid drugs have important physiological activities and are widely used clinically. They are the second largest class of drugs after antibiotics. The C1 and 2 positions of cortisone acetate are converted into prednisone acetate after introducing a double bond, the anti-inflammatory effect can be increased by 3-4 times, and the side effects caused by sodium retention can be reduced. At present, the commonly used methods in the production of steroid drugs include chemical synthesis and microbial transformation. The chemical method for ...

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Problems solved by technology

[0005] Choi et al. (1995, Journal of bacteriology 1779 (5), 4152-4156) described the gene sequence of the C1,2 position dehydrogenation gene (KsdD) in the strain of Arthrobacter Simplex, and successfully expressed this gene heterologously in Streptomyces has also studied the properties of this enzyme, but has not carried out molecular modification on Arthrobacter simplex itself, the genetic background of Arthrobacter simplex itself is unclear, and the construction of high-efficiency biotransformation Arthrobacter simplex through homologous recombination has not been reported yet

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