In Silico Analysis of Glucose Oxidase from Aspergillus niger: Potential Cysteine Mutation Sites for Enhancing Protein Stability.
In Silico Analysis of Glucose Oxidase from Aspergillus niger: Potential Cysteine Mutation Sites for Enhancing Protein Stability.
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DOI：
10.3390/bioengineering8110188
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发表时间：
2021-11-19
期刊：
影响因子：
--
通讯作者：
Jeerapan I
中科院分区：
文献类型：
--
作者：
Ittisoponpisan S;Jeerapan I
葡萄糖氧化酶（GOx）在各种应用中具有相当大的优势。然而，酶的热不稳定性仍然是一个巨大的挑战，阻碍了在控制良好的实验室之外的应用，特别是在实际的生物电子学中取得成功。人们提出了许多改进GOx以获得更好热稳定性的策略。然而，通过添加额外的二硫键来修饰这种酶还有待探索。本工作描述了黑曲霉GOx的计算机生物工程，通过对Top8000蛋白数据库中发现的二硫键特征进行分析，然后扫描适合用半胱氨酸代替的氨基酸残基对，以建立二硫键。接下来，我们从二硫键质量（键长和α角）、残基守恒的功能影响和吉布斯自由能表示的结构影响等方面预测和评估突变体GOx模型。我们发现了八个假定的残基对，它们可以形成二硫键。其中五个位于不太保守的区域，因此不太可能对功能产生有害影响。最后，两个突变，Pro149Cys和His158Cys，显示出稳定蛋白质结构的潜力，这被基于结构的稳定性分析工具证实。本研究的发现强调了使用二硫键改性作为一种新的替代技术来提高GOx的热稳定性的机会。
Glucose oxidase (GOx) holds considerable advantages for various applications. Nevertheless, the thermal instability of the enzyme remains a grand challenge, impeding the success in applications outside the well-controlled laboratories, particularly in practical bioelectronics. Many strategies to modify GOx to achieve better thermal stability have been proposed. However, modification of this enzyme by adding extra disulfide bonds is yet to be explored. This work describes the in silico bioengineering of GOx from Aspergillus niger by judiciously analyzing characteristics of disulfide bonds found in the Top8000 protein database, then scanning for amino acid residue pairs that are suitable to be replaced with cysteines in order to establish disulfide bonds. Next, we predicted and assessed the mutant GOx models in terms of disulfide bond quality (bond length and α angles), functional impact by means of residue conservation, and structural impact as indicated by Gibbs free energy. We found eight putative residue pairs that can be engineered to form disulfide bonds. Five of these are located in less conserved regions and, therefore, are unlikely to have a deleterious impact on functionality. Finally, two mutations, Pro149Cys and His158Cys, showed potential for stabilizing the protein structure as confirmed by a structure-based stability analysis tool. The findings in this study highlight the opportunity of using disulfide bond modification as a new alternative technique to enhance the thermal stability of GOx.
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影响因子：
5
作者：
Mano, Nicolas
通讯作者：
Mano, Nicolas
影响因子：
2.9
作者：
Krivov, Georgii G.;Shapovalov, Maxim V.;Dunbrack, Roland L., Jr.
通讯作者：
Dunbrack, Roland L., Jr.
影响因子：
3
作者：
Marabotti A;Del Prete E;Scafuri B;Facchiano A
通讯作者：
Facchiano A
DOI：
10.1093/protein/7.5.613
发表时间：
1994-05-01
期刊：
PROTEIN ENGINEERING
影响因子：
--
作者：
HUBBARD, SJ;GROSS, KH;ARGOS, P
通讯作者：
ARGOS, P
影响因子：
3.5
作者：
Nagano, N;Ota, M;Nishikawa, K
通讯作者：
Nishikawa, K