Novel polymorphic human UDP-glucuronosyltransferase 2A3: cloning, functional characterization of enzyme variants, comparative tissue expression, and gene induction

Michael H Court; Suwagmani Hazarika; Soundararajan Krishnaswamy; Moshe Finel; J Andrew Williams

doi:10.1124/mol.108.045500

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Novel polymorphic human UDP-glucuronosyltransferase 2A3: cloning, functional characterization of enzyme variants, comparative tissue expression, and gene induction

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Novel polymorphic human UDP-glucuronosyltransferase 2A3: cloning, functional characterization of enzyme variants, comparative tissue expression, and gene induction

Michael H Court, Suwagmani Hazarika, Soundararajan Krishnaswamy, Moshe Finel and J Andrew Williams

Molecular pharmacology, Vol.74(3), pp.744-754

09/2008

DOI: https://doi.org/10.1124/mol.108.045500

Handle:

https://hdl.handle.net/2376/100509

PMCID: PMC2574548

PMID: 18523138

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Abstract

Consensus Sequence

Organ Specificity - drug effects

Gene Expression Regulation, Enzymologic - drug effects

Liver - enzymology

Humans

Transcriptional Activation

RNA, Messenger - metabolism

Promoter Regions, Genetic - genetics

Glucuronosyltransferase - genetics

Glucuronides - metabolism

Cloning, Molecular

Recombinant Proteins - metabolism

Cell Line

RNA, Messenger - genetics

Bile Acids and Salts - metabolism

Homeodomain Proteins - genetics

Polymorphism, Genetic

Animals

Glucuronosyltransferase - metabolism

Receptors, Steroid - genetics

Deoxycholic Acid - metabolism

Hepatocyte Nuclear Factor 1 - genetics

Receptors, Glucocorticoid - genetics

Kinetics

Rifampin - pharmacology

CDX2 Transcription Factor

UDP-glucuronosyltransferases (UGTs) are critical to the detoxification of numerous drugs, environmental pollutants, and endogenous molecules. However, as yet not all of the human UGTs have been cloned and characterized. cDNA clones from the UGT2A3 gene (located on chromosome 4q13) were isolated using pooled human liver RNA. Approximately 10% of clones contained a c.1489A>G nucleotide substitution, yielding proteins with a residue 497 alanine (UGT2A3.2) instead of a threonine (UGT2A3.1). The allele frequency of this polymorphism (rs13128286) was 0.13 in a European-American population as determined by direct DNA sequencing. Of 81 structurally diverse glucuronidation substrates tested, UGT2A3 expressed by a baculovirus system selectively glucuronidated bile acids, particularly hyodeoxycholic acid at the 6-hydroxy position. Apparent K(m) values of UGT2A3.1 and UGT2A3.2 for hyodeoxycholic acid 6-glucuronidation were 69 +/- 7 and 44 +/- 12 microM, respectively. Of 29 different extrahepatic tissues evaluated by real-time polymerase chain reaction, UGT2A3 mRNA was most highly expressed in small intestine (160% of liver), colon (78% of liver), and adipose tissue (91% of liver). An in silico scan of the proximal UGT2A3 promoter/5'-regulatory region identified transcription factor consensus elements consistent with tissue-selective expression in liver (HNF1) and intestine (CXD2), as well as induction by rifampicin (pregnane X receptor). In LS180 human intestinal cells, rifampicin increased UGT2A3 mRNA by more than 4.5-fold compared with vehicle, whereas levels were not significantly affected by the arylhydrocarbon receptor ligand beta-naphthoflavone. This is the first report establishing UGT2A3 as a functional enzyme, and it represents significant progress toward the goal of having a complete set of recombinant human UGTs for comparative functional analyses.

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Title: Novel polymorphic human UDP-glucuronosyltransferase 2A3: cloning, functional characterization of enzyme variants, comparative tissue expression, and gene induction
Creators: Michael H Court - Comparative and Molecular Pharmacogenomics Laboratory, Department of Pharmacology and Experimental Therapeutics, Tufts University, 136 Harrison Ave., Boston, MA 02111, USA. michael.court@tufts.edu
Suwagmani Hazarika
Soundararajan Krishnaswamy
Moshe Finel
J Andrew Williams
Publication Details: Molecular pharmacology, Vol.74(3), pp.744-754
Academic Unit: Veterinary Clinical Sciences, Department of
Publisher: United States
Grant note: R01-GM061834 / NIGMS NIH HHS R21-GM074369 / NIGMS NIH HHS R21 GM074369-02 / NIGMS NIH HHS R21 GM074369 / NIGMS NIH HHS R01 GM061834-07 / NIGMS NIH HHS R01 GM061834 / NIGMS NIH HHS
Identifiers: 99900546513101842
Language: English
Resource Type: Journal article