Production, analysis and bioactivity of recombinant vasoactive intestinal peptide analogs.
Recombinant vasoactive intestinal polypeptide (VIP) analogs were expressed in Escherichia coli as a fusion protein containing tandemly repeated multiple copies of a synthetic VIP gene joined to glutathione S-transferase. The encoded protein contains VIP units separated by a linker peptide, potentially excisable by a double cleavage with endoprotease factor Xa and hydroxylamine. Expression of different polyVIP genes, from 1 to 32 units, was detected and the production of a 16 VIP polymer was performed. MonoVIP analogs appended by 5 or 10 amino acids at their C terminus were released by factor Xa from this polymerized product. They were then submitted to hydroxylamine cleavage to remove the linker sequence to finally obtain a recombinant VIP analog devoid of any amino acid extension. The biological activity of the recombinant polyVIP and VIP analogs was tested. Although less efficient than the natural neuropeptide, some of these components bound to VIP receptor, activated adenylate cyclase in human colonic adenocarcinoma cells and displayed a relaxation activity on guinea pig tracheal rings.
Raingeaud
J
J
Institut de Biotechnologie, Université de Limoges, France.
Lavergne
F
F
Lelievre
V
V
Muller
J M
JM
Julien
R
R
Cenatiempo
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D000242:Cyclic AMP / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics; D019073:HT29 Cells; D006801:Humans; D019811:Hydroxylamine; D006898:Hydroxylamines / Q000378:metabolism; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D011817:Rabbits; D018005:Receptors, Vasoactive Intestinal Peptide / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D014132:Trachea / Q000502:physiology; D014660:Vasoactive Intestinal Peptide / Q000031:analogs & derivatives / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification
D006898:Hydroxylamines; D009838:Oligodeoxyribonucleotides; D018005:Receptors, Vasoactive Intestinal Peptide; D011993:Recombinant Fusion Proteins; D019811:Hydroxylamine; D014660:Vasoactive Intestinal Peptide; D000242:Cyclic AMP; D005982:Glutathione Transferase