Downstream processing of recombinant hirudin produced in Saccharomyces cerevisiae

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Title
Downstream processing of recombinant hirudin produced in Saccharomyces cerevisiae
Author(s)
Bong Hyun Chung; Won Kyung Kim; J Rao; Chul Ho Kim; Sang Ki Rhee
Bibliographic Citation
Journal of Microbiology and Biotechnology, vol. 9, no. 2, pp. 179-183
Publication Year
1999
Abstract
A recombinant form of hirudin, a potent thrombin-specific inhibitor derived from the bloodsucking leech, was expressed as a secretory product in Saccharomyces cerevisiae under the control of GAL10 promoter and the mating factor α pre-pro leader sequence. In an attempt to produce recombinant hirudin (r-Hir) of therapeutic purity in large quantities, the fedbatch fermentation was carried out by using this recombinant yeast, and subsequently downstream processing was developed with the preparative-scale column chromatography systems. About 234 mg/l of biologically active r-Hir was produced as a secretory product by the fed-batch fermentation strategy developed for an efficient downstream processing. Using a two-step chromatography process (an anion exchange chromatography followed by the reverse phase HPLC), the r-Hir was purified to >98% with an overall recovery yield of 84%. According to the N-terminal amino acid sequencing, the purified r-Hir was found to have the predicted N-terminal amino acid sequence. The biological activity of the purified r-Hir to inhibit thrombin was also identical to that of the commercial hirudin.
Keyword
Recombinant hirudindownstream processingpreparative- scaleSaccharomyces cerevisiae
ISSN
1017-7825
Publisher
Korea Soc-Assoc-Inst
Type
Article
Appears in Collections:
Jeonbuk Branch Institute > Microbial Biotechnology Research Center > 1. Journal Articles
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