A new resorufin-based alpha-glucosidase assay for high-throughput screening.
A new resorufin-based alpha-glucosidase assay for high-throughput screening.
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DOI：
10.1016/j.ab.2009.04.010
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发表时间：
2009-07-01
影响因子：
2.9
通讯作者：
Zheng W
中科院分区：
文献类型：
--
作者：
Motabar O;Shi ZD;Goldin E;Liu K;Southall N;Sidransky E;Austin CP;Griffiths GL;Zheng W
α-葡萄糖苷酶的突变会导致溶酶体中糖原的积累，从而导致庞培病，一种溶酶体储存障碍。小分子伴侣分子可与酶蛋白结合，纠正突变蛋白的错误折叠和错配，成为治疗溶酶体储存障碍的一种新方法。此外，α-葡萄糖苷酶是II型糖尿病的治疗靶点，α-葡萄糖苷酶抑制剂已在临床上用作该疾病的替代治疗方法。我们开发了一种新的用于α-葡萄糖苷酶测定的荧光底物--间苯二酚α-D-吡喃葡萄糖苷。这种新底物的酶反应产物在590 nm的峰值处发射，减少了现有荧光底物4-甲基伞形花基-α-D-吡喃葡萄糖苷的荧光化合物的干扰。此外，由于该底物产物的pKa较低，因此无需添加停止液即可连续进行酶动力学测定。因此，这种新的荧光底物是一种有用的工具，用于α-葡萄糖苷酶的检测，并将促进化合物筛选，为开发庞培病的新疗法提供便利。
Mutations in α-glucosidase cause accumulation of glycogen in lysosomes resulting in Pompe disease, a lysosomal storage disorder. Small molecule chaperones that bind to enzyme proteins and correct the misfolding and mistrafficking of mutant proteins have emerged as a new therapeutic approach for the lysosomal storage disorders. In addition, α-glucosidase is a therapeutic target for type-ll diabetes, and α -glucosidase inhibitors have been used in the clinic as alternative treatments for this disease. We have developed a new fluorogenic substrate for the α-glucosidase enzyme assay, resorufin α-D-glucopyranoside. The enzyme reaction product of this new substrate emits at a peak of 590 nm, reducing the interference from fluorescent compounds seen with the existing fluorogenic substrate, 4-methylumbelliferyl-α-D-glucopyranoside. Also, the enzyme kinetic assay can be carried out continuously without the addition of stop solution, due to the lower pKa of the product of this substrate. Therefore, this new fluorogenic substrate is a useful tool for the α-glucosidase enzyme assay and will facilitate compound screening for the development of new therapies for Pompe disease.
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影响因子：
3.9
作者：
Hermans, MMP;van Leenen, D;Reuser, AJJ
通讯作者：
Reuser, AJJ
影响因子：
2.9
作者：
Coleman, Daniel J.;Studler, Missy J.;Naleway, John J.
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Mahmoud, AM;Geslevich, J;Comhaire, FH
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Comhaire, FH
影响因子：
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作者：
Parenti, Giancarlo;Zuppaldi, Alfredo;Andria, Generoso
通讯作者：
Andria, Generoso
影响因子：
15.9
作者：
REUSER, AJJ;KROOS, M;GALJAARD, H
通讯作者：
GALJAARD, H