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Molecular Characterization of the VIP Receptor Transcriptional Repressor Protein
Author(s) -
PEI LIN
Publication year - 2000
Publication title -
annals of the new york academy of sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.712
H-Index - 248
eISSN - 1749-6632
pISSN - 0077-8923
DOI - 10.1111/j.1749-6632.2000.tb06962.x
Subject(s) - repressor , dna binding domain , fusion protein , microbiology and biotechnology , transcription factor , dna binding protein , amino acid , dna , transcription (linguistics) , psychological repression , gene , biology , transcriptional regulation , reporter gene , gatad2b , peptide sequence , chemistry , biochemistry , gene expression , recombinant dna , linguistics , philosophy
A bstract : The rat type 1 VIP receptor transcriptional repressor protein (VIPR‐RP) is a recently isolated novel transcription factor. In the study reported here, the functional domains of VIPR‐RP were characterized. To map the DNA binding domain, various regions of VIPR‐RP were either transcribed and translated in vitro or expressed in and purified from E. Coli as a glutathione S‐transferase (GST) fusion. The ability of the truncated proteins to bind to VIPR‐RP specific binding sequence was tested by gel mobility shift assays. The results indicated that the amino acid sequences between 367 and 475 play an essential role for VIPR‐RP DNA binding. To determine the amino acid sequences required for transcriptional repression, fusion proteins containing the GAL4 DNA binding domain and various parts of VIPR‐RP were constructed, and their ability to repress transcription of the reporter gene containing GAL4 DNA binding sequences were tested in transiently transfected COS7 cells. The results showed that VIPR‐RP contains two separate transcriptional repression domains located between amino acids 50 to 101 and 470 to 527.