Influence of N-Terminal Domain Histidine and Proline Residues on the Substrate Selectivities of Human UDP-Glucuronosyltransferase 1A1, 1A6, 1A9, 2B7, and 2B10

Kerdpin, Oranun; Mackenzie, Peter I.; Bowalgaha, Kushari; Finel, Moshe; Miners, John O.

doi:10.1124/dmd.109.028225

Cited by 52 publications

(32 citation statements)

References 36 publications

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“…1, lane 4) that was devoid of glycosidation activity. The UGT3A2del-exon2 variant is missing all 34 amino acids of exon 2, including His35, the presumptive catalytic base required for catalysis (Kubota et al, 2007;Kerdpin et al, 2009).…”

Section: Resultsmentioning

confidence: 99%

The Novel UDP Glycosyltransferase 3A2: Cloning, Catalytic Properties, and Tissue Distribution

Mackenzie

Rogers²,

Elliot³

et al. 2011

Molecular Pharmacology

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The human UDP glycosyltransferase (UGT) 3A family is one of three families involved in the metabolism of small lipophilic compounds. Members of these families catalyze the addition of sugar residues to chemicals, which enhances their excretion from the body. The UGT1 and UGT2 family members primarily use UDP glucuronic acid to glucuronidate numerous compounds, such as steroids, bile acids, and therapeutic drugs. We showed recently that UGT3A1, the first member of the UGT3 family to be characterized, is unusual in using UDP N-acetylglucosamine as sugar donor, rather than UDP glucuronic acid or other UDP sugar nucleotides (J Biol Chem 283:36205-36210, 2008). Here, we report the cloning, expression, and characterization of UGT3A2, the second member of the UGT3 family. Like UGT3A1, UGT3A2 is inactive with UDP glucuronic acid as sugar donor. However, in contrast to UGT3A1, UGT3A2 uses both UDP glucose and UDP xylose but not UDP N-acetylglucosamine to glycosidate a broad range of substrates including 4-methylumbelliferone, 1-hydroxypyrene, bioflavones, and estrogens. It has low activity toward bile acids and androgens. UGT3A2 transcripts are found in the thymus, testis, and kidney but are barely detectable in the liver and gastrointestinal tract. The low expression of UGT3A2 in the latter, which are the main organs of drug metabolism, suggests that UGT3A2 has a more selective role in protecting the organs in which it is expressed against toxic insult rather than a more generalized role in drug metabolism. The broad substrate and novel UDP sugar specificity of UGT3A2 would be advantageous for such a function.

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Section: Resultsmentioning

confidence: 99%

The Novel UDP Glycosyltransferase 3A2: Cloning, Catalytic Properties, and Tissue Distribution

Mackenzie

Rogers²,

Elliot³

et al. 2011

Molecular Pharmacology

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“…9). It is likely that they failed to catalyze this N-glucuronidation reaction because they have a His residue at the "catalytic His" position, position 39 of the protein precursor sequence (Li and Wu, 2007), while the human UGT1A4 has a Pro residue at this position (Kerdpin et al, 2009). It may be further suggested that both medetomidine enantiomers are glucuronidated in the dog by one or two UGTs of family 2, particularly if an equivalent of the human UGT2B10 is present there (Kaivosaari et al, 2008).…”

Section: Discussionmentioning

confidence: 99%

Dog UDP-Glucuronosyltransferase Enzymes of Subfamily 1A: Cloning, Expression, and Activity

et al. 2014

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Understanding drug glucuronidation in the dog, a preclinical animal, is important but currently poorly characterized at the level of individual enzymes. We have constructed cDNAs for the 10 dog UDP-glucuronosyltransferases of subfamily 1A (dUGT1As), expressed them in insect cells, and assayed their activity as well as the activity of the nine human UGT1As, toward 14 compounds. The goal was to find out whether individual dUGT1As and individual human UGT1As have similar substrate specificities. The results revealed similarities but also many differences. For example, similarly to the human UGT1A10, dUGT1A11 exhibited high glucuronidation activity toward the 3-OH of 17-b-estradiol, 17-a-estradiol, and ethinylestradiol, and also conjugated the drug entacapone. Unlike the human UGT1A10, however, it failed to catalyze considerable rates of R-propranolol, diclofenac, and indomethacin glucuronidation.The estrogen glucuronidation assays revealed that dUGT1A8 and dUGT1A10 have a capacity to catalyze the formation of (linked) diglucuronides, an activity no human UGT1A exhibited. dUGT1A2-dUGT1A4 are homologs of the human UGT1A4, but none of them catalyzed N-glucuronidation of dexmedetomidine. Contrary to the human UGT1A4, however, dUGT1A2-dUGT1A4 catalyzed indomethacin and diclofenac glucuronidation. It may be concluded that, perhaps with the exception of UGT1A6, high similarities in substrate specificity between individual dog and human UGTs of subfamily 1A are rare or partial. Activity assays with liver and intestine microsomes of both dog and human further revealed interspecies differences, particularly in glucuronidation rates. In the dog, the microsomes assays also strongly suggested important roles for dUGTs of other subfamilies, mainly in the liver.

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“…The catalytic residues of the UGTs, a histidine at the beginning of N␣1 and an aspartate at the end of N␤4, have been addressed in several studies Patana et al, 2008;Kerdpin et al, 2009). In UGT1A1, either Asp146 or Asp151 could be the critical acid.…”

Section: Tablementioning

confidence: 99%

A Molecular Model of the Human UDP-Glucuronosyltransferase 1A1, Its Membrane Orientation, and the Interactions between Different Parts of the Enzyme

Laakkonen¹,

Finel

2010

Molecular Pharmacology

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The vertebrate UDP-glucuronosyltransferases (UGTs) are membrane-bound enzymes of the endoplasmic reticulum that process both endogenous and exogenous substrates. The human UGTs are well known biologically, but biophysical understanding is scarce, largely because of problems in purification. The one resolved crystal structure covers the C-terminal domain of the human UGT2B7. Here, we present a homology model of the complete monomeric human UGT1A1, the enzyme that catalyzes bilirubin glucuronidation. The enzyme can be seen as composed of four different domains: two large ones, the N-and C-terminal domains, and two small ones, the "envelope" helices and the transmembrane segment that includes the cytoplasmic tail. The hydrophobic core of the N-terminal domain and the two envelope helices that connect the large domains are shown to be structurally well conserved even among distant homologs and can thus be modeled with good certainty according to plant and bacterial structures. We consider alternative solutions for the highly variable N-terminal regions that probably contribute to substrate binding. The bilirubin binding site, known pathological mutations in UGT1A1, and other specific residues have been examined in the context of the model with regard to available experimental data. A putative orientation of the protein relative to the membrane has been derived from the location of predicted N-glycosylation sites. The model presents extensive interactions between the N-and C-terminal domains, the two envelope helices, and the membrane. Together, these interactions could allow for a concerted largescale conformational change during catalysis.

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Influence of N-Terminal Domain Histidine and Proline Residues on the Substrate Selectivities of Human UDP-Glucuronosyltransferase 1A1, 1A6, 1A9, 2B7, and 2B10

Cited by 52 publications

References 36 publications

The Novel UDP Glycosyltransferase 3A2: Cloning, Catalytic Properties, and Tissue Distribution

The Novel UDP Glycosyltransferase 3A2: Cloning, Catalytic Properties, and Tissue Distribution

Dog UDP-Glucuronosyltransferase Enzymes of Subfamily 1A: Cloning, Expression, and Activity

A Molecular Model of the Human UDP-Glucuronosyltransferase 1A1, Its Membrane Orientation, and the Interactions between Different Parts of the Enzyme

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