@article {10.3844/ajbbsp.2005.17.21,
article_type = {journal},
title = {Purification of A Recombinant Thrombin-like Enzyme, Gloshedobin by Egg Yolk Antibody-Coupled Adsorbents},
author = {Yang, Qing and Lei, Xuyu and Xu, Jianqiang and An, Lijia},
volume = {1},
number = {1},
year = {2005},
month = {Mar},
pages = {17-21},
doi = {10.3844/ajbbsp.2005.17.21},
url = {https://thescipub.com/abstract/ajbbsp.2005.17.21},
abstract = {The gloshedobin, a snake venom thrombin-like enzyme was biosynthesized in the soluble form and purified by egg yolk antibody-coupled adsorbents from E. coli. As His-tag is not favored from the point of view of high-level and soluble expression, we herein constructed a recombinant gloshedobin without His-tag and developed a novel egg yolk antibody (IgY)-immunoaffinity chromatography for its purification in a higher activity yield. The purification process involving Octyl Sepharose FF, IgY-immunoaffinity chromatography and Source Q, yielded 454.7U mg1 protein of interest and 34.8% activity recovery. The anti-gloshedobin IgY was obtained from eggs by injecting diluted gloshedobin into the breast muscle of laying hens and then purified by several steps including 3.5%, 8.5% and 12% polyethylene glycol-6000 precipitation and affinity chromatography using gloshedobin-coupled agarose gel. The obtained IgY was covalently linked to CNBr pre-activated agarose gel, Sepharose 4B FF, to yield immunoaffinity adsorbents. Both immunological and enzymatic activities of the purified enzyme were determined by western-blotting analysis and fibrinogen clotting assay, respectively.},
journal = {American Journal of Biochemistry and Biotechnology},
publisher = {Science Publications}
}