Expression of a recombinant porcine zona pellucida glycoprotein ZP1 in mammalian cells.
Porcine zona pellucida glycoprotein (pZP1) is a good candidate for a contraceptive vaccine. For the purpose of producing glycosylated pZP1, several types of recombinant pZP1 proteins were produced in mammalian cell lines. In the first experiment, a minigene encoding pZP1 (681 amino acids) was designed for insertion into an expression vector and then transfected to three cell lines (293T, CHO-K1, and LLC-PK1). The resulting recombinant proteins were highly glycosylated and were localized in the cytoplasm. To produce a secretory type of recombinant pZP1, in the second experiment, a cDNA coding for pZP1 excluding a putative transmembrane region and a smaller cDNA coding for 1-198 amino acid residues of pZP1 were designed to produce fusion proteins with the human IgG1 heavy chain. The resultant recombinant proteins were secreted into the supernatant from both transfected cell cultures. Recombinant secretory proteins are useful because of their simple affinity purification.
Tsubamoto
H
H
Department of Obstetrics and Gynecology, Hyogo College of Medicine, Nishinomiya, 663-8501, Japan.
Yamasaki
N
N
Hasegewa
A
A
Koyama
K
K
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D017931:DNA Primers / Q000235:genetics; D004527:Egg Proteins / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D005822:Genetic Vectors; D006801:Humans; D018374:LLC-PK1 Cells; D008562:Membrane Glycoproteins / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification; D010957:Plasmids / Q000235:genetics; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D013552:Swine; D014162:Transfection; D015044:Zona Pellucida / Q000378:metabolism*; D000072216:Zona Pellucida Glycoproteins
D017931:DNA Primers; D004527:Egg Proteins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C000606514:ZP1 protein, human; D000072216:Zona Pellucida Glycoproteins