Impact of retinal disease-associated RPE65 mutations on retinoid isomerization.
Impact of retinal disease-associated RPE65 mutations on retinoid isomerization.
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DOI：
10.1021/bi800905v
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发表时间：
2008-09-16
期刊：
影响因子：
2.9
通讯作者：
Palczewski, Krzysztof
中科院分区：
文献类型：
--
作者：
Bereta, Grzegorz;Kiser, Philip D.;Golczak, Marcin;Sun, Wenyu;Heon, Elise;Saperstein, David A.;Palczewski, Krzysztof
RPE 65基因中的致病性突变与人类的一系列先天性致盲疾病相关。我们通过对36例Leber先天性黑蒙（LCA）、62例常染色体隐性视网膜色素变性（arRP）和21例常染色体显性/隐性视锥-视杆细胞营养不良（CORD）患者的DNA直接测序，评估RPE 65基因启动子区、编码区和外显子/内含子连接处的变化。发现了15种不同的变体，其中6种是新的。有趣的是Gly 244 Val，一个靠近催化中心的新突变。为了评估该突变在RPE 65失活中的作用，我们对该突变体进行了详细的生化研究沿着并对已知对RPE 65依赖性维甲酸异构化重要的氨基酸的244个氨基酸位置进行了结构分析。RPE 65 Gly 244 Val突变体和增强型绿色荧光蛋白（EGFP）的双顺反子质粒表达使我们能够通过细胞分选和免疫印迹方法记录其在培养细胞中的不稳定性，并通过酶促测定记录其RPE 65依赖性异构酶活性的丧失。通过使用来自集胞藻属（PDB登录代码2BIW）的类胡萝卜素加氧酶（ACO）作为结构模板来构建RPE 65同源性模型，并在该模型中定位所有已知的失活突变（包括Gly 244 Val），获得了对RPE 65对类维生素A异构化的结构要求的进一步了解。
Pathogenic mutations in the RPE65 gene are associated with a spectrum of congenital blinding diseases in humans. We evaluated changes in the promoter region, coding regions, and exon/intron junctions of the RPE65 gene by direct sequencing of DNA from 36 patients affected with Leber's congenital amaurosis (LCA), 62 with autosomal recessive retinitis pigmentosa (arRP), and 21 with autosomal dominant/recessive cone-rod dystrophies (CORD). Fifteen different variants were found, of which 6 were novel. Interesting was Gly244Val, a novel mutation close to the catalytic center. To assess the role of this mutation in RPE65 inactivation, we performed detailed biochemical studies of the mutant along with a structural analysis of the 244 amino acid position with respect to amino acids known to be important for RPE65-dependent retinoid isomerization. Bicistronic plasmid expression of the RPE65 Gly244Val mutant and enhanced green fluorescent protein (EGFP) allowed us to document both its instability in cultured cells by cell sorting and immunoblotting methodology and its loss of RPE65-dependent isomerase activity by enzymatic assays. Further insights into the structural requirements for retinoid isomerization by RPE65 were obtained by using the carotenoid oxygenase (ACO) from Synechocystis (PDB accession code 2BIW) as a structural template to construct a RPE65 homology model and locating all known inactivating mutations including Gly244Val within this model.
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DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
30.8
作者：
Janecke, AR;Thompson, DA;Gal, A
通讯作者：
Gal, A
DOI：
10.1107/s0907444998003254
发表时间：
1998-09-01
期刊：
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY
影响因子：
--
作者：
Brunger, AT;Adams, PD;Warren, GL
通讯作者：
Warren, GL
DOI：
10.1007/s00417-005-0096-2
发表时间：
2006-09-01
影响因子：
2.7
作者：
El Matri, Leila;Ambresin, Aude;Munier, Francis L.
通讯作者：
Munier, Francis L.
DOI：
10.1073/pnas.0503460102
发表时间：
2005-08-30
影响因子：
11.1
作者：
Moiseyev, G;Chen, Y;Ma, JX
通讯作者：
Ma, JX