A sequence variation (I148M) in PNPLA3 associated with nonalcoholic fatty liver disease disrupts triglyceride hydrolysis.
A sequence variation (I148M) in PNPLA3 associated with nonalcoholic fatty liver disease disrupts triglyceride hydrolysis.
复制标题
DOI：
10.1074/jbc.m109.064501
复制
发表时间：
2010-02-26
期刊：
影响因子：
--
通讯作者：
Hobbs HH
中科院分区：
文献类型：
--
作者：
He S;McPhaul C;Li JZ;Garuti R;Kinch L;Grishin NV;Cohen JC;Hobbs HH
肥胖和胰岛素抵抗与甘油三酯在脂肪组织以外的组织中的沉积有关。先前，我们发现PNPLA 3（含马铃薯糖样磷脂酶结构域3蛋白）中的错义突变（I148 M）与人类肝脏甘油三酯含量增加相关。在这里，我们研究了I148 M取代对PNPLA 3的酶活性和细胞定位的影响。结构建模预测，取代蛋氨酸的异亮氨酸在残基148将限制访问的底物的催化丝氨酸在残基47。使用从Sf 9细胞部分纯化的重组PNPLA 3的体外测定证实野生型酶水解乳化甘油三酯，并且I148 M取代消除了该活性。在培养的肝细胞或小鼠肝脏中表达PNPLA 3-I148 M而非野生型PNPLA 3增加了细胞甘油三酯含量。细胞分级分离研究表明，约90%的野生型PNPLA 3分配之间的膜和脂滴，取代蛋氨酸在位置148的异亮氨酸没有改变亚细胞分布的蛋白质。这些数据与PNPLA 3-I148 M通过限制甘油三酯水解促进甘油三酯蓄积一致。
Obesity and insulin resistance are associated with deposition of triglycerides in tissues other than adipose tissue. Previously, we showed that a missense mutation (I148M) in PNPLA3 (patatin-like phospholipase domain-containing 3 protein) is associated with increased hepatic triglyceride content in humans. Here we examined the effect of the I148M substitution on the enzymatic activity and cellular location of PNPLA3. Structural modeling predicted that the substitution of methionine for isoleucine at residue 148 would restrict access of substrate to the catalytic serine at residue 47. In vitro assays using recombinant PNPLA3 partially purified from Sf9 cells confirmed that the wild type enzyme hydrolyzes emulsified triglyceride and that the I148M substitution abolishes this activity. Expression of PNPLA3-I148M, but not wild type PNPLA3, in cultured hepatocytes or in the livers of mice increased cellular triglyceride content. Cell fractionation studies revealed that ∼90% of wild type PNPLA3 partitioned between membranes and lipid droplets; substitution of isoleucine for methionine at position 148 did not alter the subcellular distribution of the protein. These data are consistent with PNPLA3-I148M promoting triglyceride accumulation by limiting triglyceride hydrolysis.