Dissociation of affinity and efficacy in KOR-3 chimeras Journal Article
| Authors: | Pan, Y. X.; Xu, J.; Ryan-Moro, J.; Mathis, J.; Hom, J. S. H.; Jianfeng, M.; Pasternak, G. W. |
| Article Title: | Dissociation of affinity and efficacy in KOR-3 chimeras |
| Abstract: | KOR-3 chimeras were constructed in which the first coding exon of KOR-3 was exchanged for the corresponding first coding exon of either MOR-1 (MOR-1/KOR-3) or DOR-1 (DOR-1/KOR3). All three clones were expressed in CHO cells and characterized with regards to their binding profiles for orphanin FQ/nociceptin (OFQ/N) and a variety of opioids as well as their functional activities in cyclase studies, 125I[Tyr14]OFQ/N labels both KOR-3 (K(D) 37 pM) and the MOR-1/KOR-3 chimera (K(D) 39 pM) equally well. Although its affinity for the DOR-1/KOR-3 chimera is quite good (K(D) 135 pM), it is slightly lower than the other two. Competition studies confirm the high affinity of OFQ/N for all three clones. However, several competitors clearly distinguish the chimeras from KOR-3, OFQ/N(1-11) competes KOR-3 (K(i) 55 nM) over 6-fold more potently than either of the chimeras (K(i) values > 350 nM). Conversely, the modest affinity of naloxone benzoylhydrazone for KOR-3 (310 nM) is greatly increased in both the MOR-1/KOR-3 (K(i) 69 nM and DOR-1/KOR-3 (K(i) 74 nM) chimeras. The remainder of the opioids tested have no appreciable affinity against any of the clones. Functionally, OFQ/N inhibits forskolin-stimulated cAMP accumulation in both the KOR-3 and the MOR-1/KOR-3 chimera by almost 40%, with IC50 values in the low nanomolar range. Little activity is seen against the DOR-1/KOR-3 chimera. Naloxone benzoylhydrazone inhibits cAMP accumulation in the KOR-3 and the DOR-1/KOR-3 chimera. Although naloxone benzoylhydrazone has higher affinity for the MOR-1/KOR-3 chimera in binding studies than KOR-3 itself, it is inactive in cyclase studies using the MOR-1/KOR-3 chimera, implying that the replacement of the first coding exon increases affinity while decreasing intrinsic activity. |
| Keywords: | controlled study; exon; nonhuman; binding affinity; animal cell; gene expression; animalia; chimera; molecular cloning; cyclic amp; protein family; morphine sulfate; opiate receptor; enkephalin[2,5 dextro penicillamine]; gene construct; adenylate cyclase; opioid receptor; cho cell; 3,4 dichloro n methyl n [2 (1 pyrrolidinyl)cyclohexyl]benzeneacetamide methanesulfonate; naloxone benzoylhydrazone; nociceptin; priority journal; article; diprenorphine; orphanin fq; kor-3; olr1 |
| Journal Title: | FEBS Letters |
| Volume: | 395 |
| Issue: | 2-3 |
| ISSN: | 0014-5793 |
| Publisher: | Wiley Blackwell |
| Date Published: | 1996-10-21 |
| Start Page: | 207 |
| End Page: | 210 |
| Language: | English |
| DOI: | 10.1016/0014-5793(96)01023-x |
| PUBMED: | 8898097 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | Article -- Export Date: 22 November 2017 -- Source: Scopus |
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