[Study on the expression of pig zona pellucida-3 beta excluding N-terminus signal peptide and C-terminus transmembrance-like domain in Escherichia coli].
Pig oocytes are surrounded by an acellular translucent envelope-zona pellucida(pZP), which is comprised of three biochemically and immunologically distinct glycoproteins(pZP1, pZP3 alpha and pZP3 beta). Due to the cross-reactivity of anti-pZP3 beta antibodies with human ZP in vitro, pZP3 beta has been considered as potential one of immunogens for developing human contraceptive vaccine. In order to express pZP3 beta directly in E. coli; we amplified the core fragment of pZP3 beta cDNA by PCR that was deleted 5'- and 3'-terminal sequences of coding for signal peptide and transmembrane-like domain. The DNA sequenced EcoRI and SalI restriced core fragment was cloned in a frame downstream of PRPL promoter in the pBV221 vector. SDS-PAGE analysis showed that pZP3 beta protein was expressed especially in E. coli after thermal induction, which the expression band displayed the molecular weight of about 38 kD matching with its deduced molecular weight(36.5 kD). In addition, the specially expressed protein band on SDS-PAGE gel showed specific immunological reaction with anti-pig ZP IgGs of rabbit in Western blot.
Xu
W X
WX
Shanghai Institute of Planned Parenthood Research, Shanghai 200032, China.
Qiu
D Y
DY
Xiong
Y
Y
Sheng
Q X
QX
Gu
S H
SH
Cao
G T
GT
Ying
K
K
Xie
Y
Y
D000818:Animals; D004527:Egg Proteins / Q000096:biosynthesis / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics*; D010957:Plasmids; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins / Q000096:biosynthesis*; D013552:Swine; D000072216:Zona Pellucida Glycoproteins
D004527:Egg Proteins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D000072216:Zona Pellucida Glycoproteins