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Glucuronidation of D-Luciferin In Vitro: Isoform Selectivity and Kinetics Characterization
1School of Life Science and Medicine, Dalian University of Technology, Panjin, China. xiayl@dlut.edu.cn.
Background:
D-luciferin is one of the most commonly used substrates in bioluminescence imaging for real-time monitoring of sophisticated biological processes in models of human biology or disease in vitro and in vivo. D-luciferin is rapidly cleared from blood circulation after being exogenously delivered in vivo and the presence of phenolic groups indicates that glucuronide conjugation is a possible metabolic pathway for the compound.
Objectives:
This study aimed to characterize the glucuronidation pathway of D-luciferin in human liver microsomes (HLM) and human intestine microsomes (HIM).
Methods:
HLM and HIM were employed to catalyze the glucuronidation of D-luciferin in vitro. The activity of recombinant uridine-diphospho-glucuronosyltransferase (UGT) isoforms towards D-luciferin glucuronidation was screened. Chemical inhibition assay and kinetic analysis was combined to determine the UGT isoforms responsible for the formation of D-luciferin glucuronide in HLM and HIM.
Results:
D-luciferin could be catalyzed to form one mono-glucuronide which was characterized as 6'-O-glucuronide in HLM and HIM. UGT1A1, 1A3, 1A6, 1A8, 1A9 and 1A10 participated in the formation of D-luciferin glucuronide, with UGT1A1 exhibiting the highest catalytic activity. Both chemical inhibition assays and kinetic analysis showed that UGT1A1 and UGT1A3 played important roles in D-luciferin-6'-O-glucuronidation in HLM and HIM, with UGT1A6 also giving a non-negligible contribution to this biotransformation in HLM.
Conclusions:
UGT1A1, UGT1A3 and UGT1A6 were responsible for 6'-O-glucuronidation of D-luciferin in HLM, while UGT1A1 and UGT1A3 were the major contributors to this biotransformation in HIM.
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