Replacement of Serine363and Serine375Codons by Alanine in Ratμ-Opioid ReceptorcDNA Replacement of serine363and serine375by alanine in μ-receptor
Iranian Journal of Pharmaceutical Sciences,
Vol. 3 No. 1 (2007),
15 January 2007
,
Page 13-18
https://doi.org/10.22037/ijps.v3.40328
Abstract
The aim of this study was to use site directed mutagenesis technique to constructa vector in which serine363and serine375residues of the COOH-terminal portion ofthe μ-opioid receptor (MOR) were substituted by alanine. These constructs areessential in studying G-protein coupled receptor kinase-mediated MOR desensiti-zation. The nested PCR carried out for conversion of serine363and serine375to alanineresulted in the production of a band comparable to the expected size of 1400 bp.Restriction analysis of these bands confirmed the integrity of the PCR products.Ligation of the mutated PCR product into pcDNA3 and its digestion with appropriaterestriction enzymes further confirmed the integrity of the PCR product and itsorientation into the vector.
- Opioid receptor
- pcDNA3
- Serine363
- Serine375
How to Cite
References
[2]Nestler EJ. Under siege: The brain on opiates.Neuron1996; 16: 897-900.
[3]Nestler EJ, Aghajanian GK. Molecular andcellular basis of addiction. Science1997; 278: 58-63.
[4]Pitcher JA, Freedman NJ, Lefkowitz RJ. G-protein-coupled receptor kinases. Annu RevBiochem1998; 67: 653-92.
[5]Wang HL, Chang WT, Hsu CY, Huang PC, ChowYW, Li AH. Identification of two C-terminalamino acids, Ser(355) and Thr(357), required forshort-term homologous desensitization of mu-opioid receptors. Biochem Pharmacol2002; 64:257-66.
[6]Reisine T, Bell GI. Molecular biology of opioidreceptors. Trends Neurosci 1993; 16: 506-10.
[7]Zaki PA, Bilsky EJ, Vanderah TW, Lai J, EvansCJ, Porreca F. Opioid receptor types and subtypes:The delta receptor as a model. Annu RevPharmacol Toxicol1996; 36: 379-401.
[8]Gudermann T, Schoneberg T, Schultz G.Functional and structural complexity of signaltransduction via G-protein-coupled receptors.Annu Rev Neurosci 1997; 20: 399-427.
[9]Savarese TM, Fraser CM. In vitromutagenesisand the search for structure-function relationshipsamong G-protein-coupled receptors. Biochem J1992; 283 ( Pt 1): 1-19.
[10]Strader CD, Fong TM, Tota MR, Underwood D,Dixon RA. Structure and function of G- protein-coupled receptors. Annu Rev Biochem 1994; 63:101-32.
[11]El Kouhen R, Burd AL, Erickson-Herbrandson LJ,Chang CY, Law PY, Loh HH. Phosphorylation ofSer363, Thr370, and Ser375 residues within thecarboxyl tail differentially regulates mu-opioidreceptor internalization. J Biol Chem2001; 276:12774-80.
[12]Schulz S, Mayer D, Pfeiffer M, Stumm R, KochT, Hollt V. Morphine induces terminal micro-opioid receptor desensitization by sustainedphosphorylation of serine-375. EMBO J2004; 23:3282-9.
[13]Zimprich A, Simon T, Hollt V. Transfected rat muopioid receptors (rMOR1 and rMOR1B) stimulatephospholipase C and Ca2+mobilization.Neuroreport1995; 7: 54-6.
[14]Sambrook J, Russel DW, Sambrook J. Molecularcloning: Alaboratory manual. New York: ColdSpring Harbor Laboratory Press, 2002.
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