Abstract
UDP-Glucuronosyltransferases UGT2B2I and UGT2B22 cDNA which encode guinea pig UGT55K and UGT59K have been cloned and characterized. It has been suggested that an active glucuronide of morphine, morphine 6-glucuronide (M6G), is responsible for the analgesia in humans. Besides humans, guinea pigs have the ability to glucuronidate morphine to M6G with great efficiency. The purification of morphine UGT in this animal species gave us a preparation of a possible hetero-oligomer of UGT55K and UGT59K. The purpose of the present work was to clone the isoforms and characterize the homo and hetero-oligomers. UGT2B21 and UGT2B22 cDNAs were cloned by screening of the Hartley guienea pig liver cDNA libraries. UGT2B2I cDNA containing full length of open reading frame were obtained by 5' and 3'-RACE, and further analysis of the full length RT-PCR product of 1821 by encoding 528 amino acids. UGT2B22 cDNA which was containing length of open reading frame was obtained by 5'-RACE, was 2508 by and deduced amino acids were 529. UGT2B21 and UGT2B22 expression vectors were constructed in pSVL. COS-1 cells were transfected with UGT2B21 and/or UGT2B22 expression vector by Trans-ITTM, and the UGT activity in the COS I cell microsomes was determined. UGT2B21 glucuronidated the morphine 3-position, 4-hydroxybiphenyl, bomeol, testosterone, androsterone and estriol. M6G activity and chloramphenicol glucuronidation were only seen in the microsomes of co-expressed UGT2B2I and UGT2B22. As yet, we know of no substrate for UGT2B22. The present results indicate that UGT isoforms could act as hetero-oligomers by accessing a broader range of substrates than homo-oligomers.
