Homo- and hetero-dimerization of human UDP-glucuronosyltransferase 2B7 (UGT2B7) wild type and its allelic variants affect zidovudine glucuronidation activity.
Homo- and hetero-dimerization of human UDP-glucuronosyltransferase 2B7 (UGT2B7) wild type and its allelic variants affect zidovudine glucuronidation activity.
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DOI：
10.1016/j.bcp.2015.03.002
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发表时间：
2015-05
影响因子：
5.8
通讯作者：
Lingmin Yuan;Sainan Qian;Yongsheng Xiao;Hong-ying Sun;S. Zeng
中科院分区：
文献类型：
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作者：
Lingmin Yuan;Sainan Qian;Yongsheng Xiao;Hong-ying Sun;S. Zeng
大多数人UDP-葡萄糖醛酸基转移酶(UGT；EC 2.4.1.17)基因含有导致氨基酸替换的非同义单核苷酸多态(NsSNPs)。由nsSNPs引起的等位基因变异可能表现出酶活性缺失或降低。UGT2B7是最重要的UGT之一，可糖醛酸化丰富的内生和外生物质，如雌三醇、吗啡和抗癌药物。在UGT2B7基因中发现了3个nsSNP，分别为UGT2B7*71S(211G&gT；T)、UGT2B7*2(802C>；T)和UGT2B7*5(1192G>；A)，它们编码等位基因UGT2B7*71S(A71S)、UGT2B7*2(H268Y)和UGT2B7*5(D398N)。已观察到UGT2B7形成影响其酶活性的寡聚体，在本研究中，我们通过系统的定量荧光共振能量转移(FRET)分析并结合免疫共沉淀实验，研究了UGT2B7野生型(WT)、A71S、H268Y和D398N等位基因之间的蛋白质-蛋白质相互作用。定量FRET分析表明，UGT2B7等位酶形成同源和异源二聚体，并在供体-受体距离上表现出明显的特征。N-末端结构域上的密码子71和密码子268都参与了二聚体相互作用。免疫共沉淀实验也证明了UGT2B7等位酶形成了稳定的二聚体。以齐多夫定为底物，进一步考察了同二聚体和杂二聚体的葡萄糖醛酸化活性。H268Y和D398N均通过形成杂二聚体来抑制WT和A71S的活性，而当WT与A71S异二聚体形成时，WT和A71S的活性增加。此外，齐多夫定葡萄糖醛酸化活性与FRET距离有关。这些发现为深入了解UGT2B7中氨基酸替换对齐多夫定葡萄糖醛酸化反应的影响，以及蛋白质-蛋白质相互作用与葡萄糖醛酸化活性之间的关联提供了见解。
Most humanUDP-glucuronosyltransferase(UGT; EC 2.4.1.17) genes contain non-synonymous single nucleotide polymorphisms (nsSNPs) which cause amino acid substitutions. Allelic variants caused by nsSNPs may exhibit absent or reduced enzyme activity. UGT2B7 is one of the most important UGTs that glucuronidates abundant endobiotics and xenobiotics, such as estriol, morphine, and anticancer drugs. Three nsSNPs,UGT2B7*71S(211G>T),UGT2B7*2(802C>T) andUGT2B7*5(1192G>A) are observed in theUGT2B7gene, and they code for allozymes UGT2B7*71S (A71S), UGT2B7*2 (H268Y), and UGT2B7*5 (D398N). UGT2B7 has been observed to form oligomers that affect its enzymatic activity and in this study, we investigated protein–protein interactions among UGT2B7 allozymes wild type (WT), A71S, H268Y and D398N, by performing a systematic quantitative fluorescence resonance energy transfer (FRET) analysis in combination with co-immunoprecipitation assay. Quantitative FRET analysis revealed that UGT2B7 allozymes formed homo- and hetero-dimers and showed distinct features in donor–acceptor distances. Both codon 71 and codon 268 in the N-terminal domain were involved in the dimeric interaction. Co-immunoprecipitation experiments also proved that UGT2B7 allozymes formed stable dimers. The glucuronidation activities of homo- and hetero-dimers were further tested with zidovudine as the substrate. An increase in activity was observed when WT hetero-dimerized with A71S compared with homo-dimers, while both H268Y and D398N impaired the activity of WT and A71S by forming hetero-dimers. In addition, zidovudine glucuronidation activity is associated with FRET distance. These findings provide insights into the consequences of amino acid substitution in UGT2B7 on zidovudine glucuronidation and the association between protein–protein interaction and glucuronidation activity.