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Title 

Downstream processing of recombinant hirudin produced in Saccharomyces cerevisiae

Authors 

Bong Hyun ChungWon Kyung KimJ RaoChul Ho KimSang Ki Rhee

Publisher 

The Korean Society for Applied Microbiology

Issue Date 

1999

Citation 

Journal of Microbiology and Biotechnology, vol. 9, no. 2, pp. 179-183

Keywords 

Recombinant hirudindownstream processingpreparative- scaleSaccharomyces cerevisiae

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.

ISSN 

1017-7825

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2017-04-19


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