| Title: | Importance of asparagine residues at positions 13 and 26 on the amino-terminal domain of human somatostatin receptor subtype-5 in signalling |
| Author(s): | Togawa S; Ishii J; Ishikura A; Tanaka T; Ogino C; Kondo A; |
| Address: | "Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, Kobe, Japan" |
| ISSN/ISBN: | 1756-2651 (Electronic) 0021-924X (Linking) |
| Abstract: | "N-linked oligosaccharides or asparagine residues are often involved in G protein-coupled receptor functions. Focusing on Asn13 and Asn26 positioned on N-linked glycosylation motifs in the amino-terminal domain of human somatostatin receptor subtype-5 (hSSTR5), we performed site-directed mutagenesis and evaluated the mutants by using yeast cells as the host strain. This is because analysing the complicated signalling in mammalian cell lines is simplified by the utilization of the monopolistic pheromone signalling pathway in yeast. Western blot analysis and confocal laser scanning microscope observation showed that Asn13 and/or Asn26 mutations had no effects on cell-surface expression of hSSTR5 in yeast. By using an engineered yeast strain of Saccharomyces cerevisiae, which induces the expression of the green fluorescent protein (GFP) reporter gene in response to the agonist-specific signal transduction, it was demonstrated that a single mutation of two asparagine residues attenuated the somatostatin-specific signalling levels, and the double mutant significantly lost the signalling ability. These results clearly show the importance of these asparagine residues in the agonist-specific signalling of hSSTR5, although it was not enough to identify the consequence of oligosaccharides" |
| Keywords: | "Amino Acid Substitution Asparagine Blotting, Western Glycosylation Green Fluorescent Proteins Humans Membrane Glycoproteins/*chemistry/metabolism Microscopy, Confocal Mutagenesis, Site-Directed Receptors, G-Protein-Coupled/*chemistry/metabolism *Receptors;" |
| Notes: | "MedlineTogawa, Shota Ishii, Jun Ishikura, Atsushi Tanaka, Tsutomu Ogino, Chiaki Kondo, Akihiko eng Research Support, Non-U.S. Gov't England 2010/03/09 J Biochem. 2010 Jun; 147(6):867-73. doi: 10.1093/jb/mvq022. Epub 2010 Mar 5" |
