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Expression and characterization of VIP and two VIP mutants in NIH 3T3 cells
Author(s) -
Wulff Birgitte S.,
Georg Birgitte,
Fahrenkrug Jan
Publication year - 1994
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(94)80237-8
Subject(s) - vasoactive intestinal peptide , furin , receptor , pancreatic polypeptide , cell culture , biochemistry , biology , mutant , peptide , enteroendocrine cell , adenylate kinase , microbiology and biotechnology , cyclase , chemistry , enzyme , hormone , neuropeptide , gene , endocrine system , glucagon , genetics
Prepro‐vasoactive intestinal peptide (preproVIP) was expressed in NIH 3T3 cells, and the preproVIP‐derived peptides produced by the cells were analyzed by chromatography combined with sequence‐specific radio‐immunoanalysis. In accordance with what has previously been reported on processing in non‐endocrine cell lines, the VIP precursor was processed poorly in these non‐endocrine cells. Mainly an extended form of VIP could be detected in the media from the cells, and no immunoreactivity specific for amidated VIP was found. However, by changing the dibasic cleavage site positioned N‐terminal to the VIP sequence in the precursor into the consensus sequence (Arg,X,Lys/Arg,Arg) for the ubiquitous processing enzyme furin, thought to process, e.g. insulin receptors, factor VII, and by deleting residues 156–170 in the VIP precursor, expression of amidated VIP was obtained in this fibroblast cell line. Peptides from the wild‐type VIP precursor as well as peptides from the mutated VIP precursor were found to be able to stimulate the adenylate cyclase in cells expressing the VIP receptor.