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ISSN 0582-9879 Acta Biochim et Biophysica Sinica 2004, 36(5):331-335 CN
31-1300/Q
Expression and Identification
of Mutated Osteoprotegerin in Culture Cells and
Larvae of Silkworm
Hai-Long XIAO, Yong-Feng JIN, and Yao-Zhou ZHANG1*
( Institute of Biochemistry, College of Life Sciences, Zhejiang University,
Hangzhou 310029, China;
1College of Life Sciences, Zhejiang University of Science &
Technology, Hangzhou 310018, China )
Abstract The mutated osteoprotegerin (OPG-372) gene was inserted into the baculovirus transfer vector pBacPAK8, and the recombinant plasmid was co-transfected with linearized Bm-BacPAK6 virus DNA into BmN cells, then homologous recombination occurred inside the cells. The recombinant virus BmNPV-OPG-372 was screened and identified by Southern blotting. The recombinant human OPG-372 was expressed in cultured cells and the larvae of silkworm by inoculation of recombinant virus. The expression products were run on the SDS-PAGE and their immunoreactivities were determined by Western blotting. It was found that a 42 kD recombinant protein was expressed in BmN cells and a 46 kD one in larvae respectively. The bioactivities of the recombinant proteins were determined by hypocalcemic effect assay in the mice. The results showed that the recombinant proteins had a significant hypocalcemic effect on mice sera.
Key words OPG-372; silkworm baculovirus expression system; hypocalcemic effect
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Received: January 7,
2004 Accepted: March 31, 2004
*Corresponding author: Tel, 86-571-86843190; Fax, 86-571-86843198; E-mail, yaozhou@chinagene.com