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Expression of recombinant Hirudin in transgenic mice milk driven by the goat β‐casein promoter

Expression of recombinant Hirudin in transgenic mice milk driven by the goat β‐casein promoter Hirudin, isolated from the leech Hirudo medicinalis, inhibits thrombin directly and several expression systems have been used to produce recombinant Hirudin (rHirudin) for pharmaceutical purposes. A DNA fragment containing the Hirudin coding sequence and goat β‐casein secretion signal was chemically synthesized in this study. The synthetic DNA then was further constructed into a goat β‐casein expression vector for mouse transgenesis. Four lines of transgenic mice were successfully developed and one line showed a meaningful anti‐thrombin activity of 40,000 anti‐thrombin units (ATU)/mL in their milk. In this animal line, Hirudin mRNA was found in samples of uterus and kidney with insignificant anti‐thrombin activity (≤ 280 ATU/g wet tissue); however, mammary glands showed a higher activity of 780 ATU/g wet tissue. Transgenic mice showed no evident physical abnormality. The purified rHirudin was further analyzed by amino acid analysis and was found to contain a tyrosine O‐sulfate residue that is absent in rHirudin expression either through Escherichia coli or yeast host systems. Experimental results demonstrated that the β‐casein‐promoted Hirudin transgene could be successfully expressed in a murine model and may be applicable to large mammals such as livestock for mass production of rHirudin for pharmaceuticals. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Biotechnology Journal Wiley

Expression of recombinant Hirudin in transgenic mice milk driven by the goat β‐casein promoter

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References (43)

Publisher
Wiley
Copyright
Copyright © 2008 Wiley Subscription Services, Inc., A Wiley Company
ISSN
1860-6768
eISSN
1860-7314
DOI
10.1002/biot.200800069
pmid
18543243
Publisher site
See Article on Publisher Site

Abstract

Hirudin, isolated from the leech Hirudo medicinalis, inhibits thrombin directly and several expression systems have been used to produce recombinant Hirudin (rHirudin) for pharmaceutical purposes. A DNA fragment containing the Hirudin coding sequence and goat β‐casein secretion signal was chemically synthesized in this study. The synthetic DNA then was further constructed into a goat β‐casein expression vector for mouse transgenesis. Four lines of transgenic mice were successfully developed and one line showed a meaningful anti‐thrombin activity of 40,000 anti‐thrombin units (ATU)/mL in their milk. In this animal line, Hirudin mRNA was found in samples of uterus and kidney with insignificant anti‐thrombin activity (≤ 280 ATU/g wet tissue); however, mammary glands showed a higher activity of 780 ATU/g wet tissue. Transgenic mice showed no evident physical abnormality. The purified rHirudin was further analyzed by amino acid analysis and was found to contain a tyrosine O‐sulfate residue that is absent in rHirudin expression either through Escherichia coli or yeast host systems. Experimental results demonstrated that the β‐casein‐promoted Hirudin transgene could be successfully expressed in a murine model and may be applicable to large mammals such as livestock for mass production of rHirudin for pharmaceuticals.

Journal

Biotechnology Journal – Wiley

Published: Aug 1, 2008

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