A novel variant L263F in human inosine 5'-monophosphate dehydrogenase 2 is associated with diminished enzyme activity.
A novel variant L263F in human inosine 5'-monophosphate dehydrogenase 2 is associated with diminished enzyme activity.
复制标题
人肌苷 5-单磷酸脱氢酶 2 中的新变体 L263F 与酶活性降低有关。
DOI：
10.1097/fpc.0b013e328012b8cf
复制
发表时间：
2007
影响因子：
2.6
通讯作者：
Burckart,GilbertJ
中科院分区：
文献类型：
--
作者：
Wang,Jian;Zeevi,Adriana;Webber,Steve;Girnita,DianaM;Addonizio,Linda;Selby,Rick;Hutchinson,IanV;Burckart,GilbertJ
方法对152例实体器官移植患者进行了IMPDH2基因外显子和外显子/内含子交界处的DNA筛查，并进行了双向DNA测序。利用定点突变技术构建基因突变体，转化5‘-单磷酸脱氢酶缺陷型大肠杆菌H712。结果在IMPDH2基因中发现了9个基因变异，其中较为罕见的等位基因频率在0.5%~10.2%之间。鉴定出一个新的非同义突变体L263F，动力学测定表明，突变体L263F的5‘-单磷酸肌苷脱氢酶活性降低到野生型的10%。突变株L263F对霉酚酸的抑制作用与野生型相当，对肌苷5‘-单磷酸和烟酰胺腺嘌呤二核苷酸的抑制作用Km无明显变化。结论IMPDH2的遗传多样性较低，但非同义突变株L263F对肌苷5’-单磷酸脱氢酶活性有显著影响。这种新的功能变异可能是导致移植患者基线肌苷5‘-单磷酸脱氢酶活性个体间差异以及药物疗效和不良事件的因素之一。
MethodsDNA samples from 152 solid organ transplant patients were screened at exons and exon/intron junctions of the IMPDH2 genes by PCR amplification followed by bidirectional direct DNA sequencing. Genetic variant was constructed by site-directed mutagenesis and transformed to an inosine 5′-monophosphate dehydrogenase-deficient strain of Escherichia coli h712. Proteins were purified to homogeneity and the enzymatic activity was measured by reduced nicotinamide adenine dinucleotide production.ResultsNine genetic variants were identified in the IMPDH2 gene, with frequencies of the rarer alleles ranging from 0.5 to 10.2%. A novel nonsynonymous variant L263F was identified, and the kinetic assay demonstrated that the inosine 5′-monophosphate dehydrogenase activity of L263F variant was decreased to 10% of the wild-type. The K i for mycophenolic acid inhibition of the L263F variant was comparable with the wild-type, and the variant K m for inosine 5′-monophosphate and nicotinamide adenine dinucleotide did not change significantly.ConclusionsIMPDH2 has low genetic diversity, but the nonsynonymous variant L263F has a significant impact on inosine 5′-monophosphate dehydrogenase activity. This novel functional variant may be one of the factors contributing to the inter-individual difference of baseline inosine 5′-monophosphate dehydrogenase activity as well as drug efficacy and adverse events in transplant patients.