Crystal structure of a mutant glycosylasparaginase shedding light on aspartylglycosaminuria-causing mechanism as well as on hydrolysis of non-chitobiose substrate.
Crystal structure of a mutant glycosylasparaginase shedding light on aspartylglycosaminuria-causing mechanism as well as on hydrolysis of non-chitobiose substrate.
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DOI：
10.1016/j.ymgme.2017.04.008
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发表时间：
2017-06
影响因子：
3.8
通讯作者：
Guo HC
中科院分区：
文献类型：
--
作者：
Pande S;Lakshminarasimhan D;Guo HC
糖基冬酰胺酶 (GA) 是一种酰胺酶，可在溶酶体中裂解 Asn 连接的糖蛋白。这种酶的缺乏会导致糖天冬酰胺在细胞溶酶体中积聚，从而导致一种称为天冬氨酰糖胺尿症 (AGU) 的遗传性疾病。为了更好地理解单残基从甘氨酸变为天冬氨酸的致病突变的机制，我们在相应位置生成了模型突变酶（命名为G172D突变体）。在这里，我们报道了成熟 G172D 突变体的 1.8Å 分辨率晶体结构，并分析了其水解酶活性低的原因。成熟 G172D 和野生型 GA 模型的比较表明，催化位点附近 Asp 172 的存在会影响成熟 G172D 中的底物分解代谢，使其底物处理效率降低。最近的研究还表明，GA 能够通过其外切水解酶活性处理缺乏壳二糖（聚糖、N-乙酰基二糖、NAcGlc）部分的底物。这种类型的催化机制尚不清楚。 L-天冬氨酸 β-异羟肟酸酯 (β-AHA) 是一种非壳二糖底物，已知与 GA 相互作用。为了研究非壳二糖底物加工的潜在机制，我们通过与先前发表的与 β-AHA 分子复合的 G172D 突变体前体进行比较，构建了 GA-β-AHA 复合物结构。基于这个复杂的模型，提出了 GA 水解 β-AHA 的机理。
Glycosylasparaginase (GA) is an amidase that cleaves Asn-linked glycoproteins in lysosomes. Deficiency of this enzyme causes accumulation of glycoasparagines in lysosomes of cells, resulting in a genetic condition called aspartylglycosaminuria (AGU). To better understand the mechanism of a disease-causing mutation with a single residue change from a glycine to an aspartic acid, we generated a model mutant enzyme at the corresponding position (named G172D mutant). Here we report a 1.8Å resolution crystal structure of mature G172D mutant and analyzed the reason behind its low hydrolase activity. Comparison of mature G172D and wildtype GA models reveals that the presence of Asp 172 near the catalytic site affects substrate catabolism in mature G172D, making it less efficient in substrate processing. Also recent studies suggest that GA is capable of processing substrates that lack a chitobiose (Glycan, N-acetylchiobios, NAcGlc) moiety, by its exo-hydrolase activity. The mechanism for this type of catalysis is not yet clear. L-aspartic acid β-hydroxamate (β-AHA) is a non-chitobiose substrate that is known to interact with GA. To study the underlying mechanism of non-chitobiose substrate processing, we built a GA-β-AHA complex structure by comparing to a previously published G172D mutant precursor in complex with a β-AHA molecule. A hydrolysis mechanism of β-AHA by GA is proposed based on this complex 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：
4.8
作者：
Guan, CD;Cui, T;Comb, D
通讯作者：
Comb, D
影响因子：
64.5
作者：
Xu, QA;Buckley, D;Guo, HC
通讯作者：
Guo, HC
DOI：
10.1107/s0907444999011798
发表时间：
1999-11-01
期刊：
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY
影响因子：
--
作者：
Cui, T;Liao, PH;Guo, HC
通讯作者：
Guo, HC
影响因子：
5.7
作者：
Sui, Lufei;Lakshminarasimhan, Damodharan;Guo, Hwai-Chen
通讯作者：
Guo, Hwai-Chen