Fusariumgraminearum 的脱卤糖-6-磷酸磷酸酶的表达和酶特性。

IF 1.4 4区生物学 Q4 BIOCHEMICAL RESEARCH METHODS

Protein expression and purification Pub Date : 2024-11-05 DOI:10.1016/j.pep.2024.106619

Xuebiao Zhang , Le Chen , Zhong Ni , Chao Xu , Qinyan Wu , Yiqing Zhuang

{"title":"Fusariumgraminearum 的脱卤糖-6-磷酸磷酸酶的表达和酶特性。","authors":"Xuebiao Zhang , Le Chen , Zhong Ni , Chao Xu , Qinyan Wu , Yiqing Zhuang","doi":"10.1016/j.pep.2024.106619","DOIUrl":null,"url":null,"abstract":"<div><div>This study presents an exhaustive characterization of the enzymatic attributes and structural properties of trehalose-6-phosphate phosphatase (TPP) derived from <em>Fusarium graminearum</em>. Enzyme activity was evaluated through a meticulously designed enzymatic assay. The findings indicate that the molecular weight of the enzyme is approximately 99.8 kDa, with an optimal reaction temperature and pH of 40 °C and 6.5, respectively. Magnesium ions (Mg<sup>2+</sup>) markedly enhance the enzymatic activity, resulting in a specific activity of 1.795 U/μg. Kinetic analysis revealed a <em>K</em><sub>m</sub> value of 0.96 μmol/L and a <em>V</em><sub>max</sub> of 15.79 μmol/L/min. Subsequent computational analysis elucidated the three-dimensional architecture of the enzyme and identified the binding site for the substrate trehalose-6-phosphate (T6P). T6P was found to form hydrogen bonds with TPP at residues Lys754, Arg720, His665, Glu758, and Asn756. Additionally, hydrophobic interactions were observed between T6P and residues Phe802, Ile610, Asp801, Pro752, and Gly753. The binding energy calculated for the T6P-TPP complex stood at −5.7 kcal/mol.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"226 ","pages":"Article 106619"},"PeriodicalIF":1.4000,"publicationDate":"2024-11-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Trehalose-6-phosphate phosphatase expression and enzymatic properties of Fusarium graminearum\",\"authors\":\"Xuebiao Zhang , Le Chen , Zhong Ni , Chao Xu , Qinyan Wu , Yiqing Zhuang\",\"doi\":\"10.1016/j.pep.2024.106619\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><div>This study presents an exhaustive characterization of the enzymatic attributes and structural properties of trehalose-6-phosphate phosphatase (TPP) derived from <em>Fusarium graminearum</em>. Enzyme activity was evaluated through a meticulously designed enzymatic assay. The findings indicate that the molecular weight of the enzyme is approximately 99.8 kDa, with an optimal reaction temperature and pH of 40 °C and 6.5, respectively. Magnesium ions (Mg<sup>2+</sup>) markedly enhance the enzymatic activity, resulting in a specific activity of 1.795 U/μg. Kinetic analysis revealed a <em>K</em><sub>m</sub> value of 0.96 μmol/L and a <em>V</em><sub>max</sub> of 15.79 μmol/L/min. Subsequent computational analysis elucidated the three-dimensional architecture of the enzyme and identified the binding site for the substrate trehalose-6-phosphate (T6P). T6P was found to form hydrogen bonds with TPP at residues Lys754, Arg720, His665, Glu758, and Asn756. Additionally, hydrophobic interactions were observed between T6P and residues Phe802, Ile610, Asp801, Pro752, and Gly753. The binding energy calculated for the T6P-TPP complex stood at −5.7 kcal/mol.</div></div>\",\"PeriodicalId\":20757,\"journal\":{\"name\":\"Protein expression and purification\",\"volume\":\"226 \",\"pages\":\"Article 106619\"},\"PeriodicalIF\":1.4000,\"publicationDate\":\"2024-11-05\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Protein expression and purification\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S1046592824001918\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q4\",\"JCRName\":\"BIOCHEMICAL RESEARCH METHODS\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Protein expression and purification","FirstCategoryId":"99","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S1046592824001918","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}

引用次数: 0

摘要

本研究详尽描述了从禾谷镰刀菌（Fusarium graminearum）中提取的三卤糖-6-磷酸磷酸酶（TPP）的酶属性和结构特性。通过精心设计的酶测定法对酶活性进行了评估。研究结果表明，该酶的分子量约为 99.8 kDa，最佳反应温度和 pH 值分别为 40 °C 和 6.5。镁离子（Mg2+）能显著提高酶的活性，使酶的比活达到 1.795 U/μg 。动力学分析表明，Km 值为 0.96 μmol/L，Vmax 为 15.79 μmol/L/min。随后的计算分析阐明了该酶的三维结构，并确定了底物三卤糖-6-磷酸（T6P）的结合位点。研究发现，T6P 与 TPP 在 Lys754、Arg720、His665、Glu758 和 Asn756 残基上形成氢键。此外，还观察到 T6P 与残基 Phe802、Ile610、Asp801、Pro752 和 Gly753 之间的疏水相互作用。计算得出的 T6P-TPP 复合物结合能为 -5.7 kcal/mol。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

查看原文本刊更多论文

Trehalose-6-phosphate phosphatase expression and enzymatic properties of Fusarium graminearum

This study presents an exhaustive characterization of the enzymatic attributes and structural properties of trehalose-6-phosphate phosphatase (TPP) derived from Fusarium graminearum. Enzyme activity was evaluated through a meticulously designed enzymatic assay. The findings indicate that the molecular weight of the enzyme is approximately 99.8 kDa, with an optimal reaction temperature and pH of 40 °C and 6.5, respectively. Magnesium ions (Mg²⁺) markedly enhance the enzymatic activity, resulting in a specific activity of 1.795 U/μg. Kinetic analysis revealed a K_m value of 0.96 μmol/L and a V_max of 15.79 μmol/L/min. Subsequent computational analysis elucidated the three-dimensional architecture of the enzyme and identified the binding site for the substrate trehalose-6-phosphate (T6P). T6P was found to form hydrogen bonds with TPP at residues Lys754, Arg720, His665, Glu758, and Asn756. Additionally, hydrophobic interactions were observed between T6P and residues Phe802, Ile610, Asp801, Pro752, and Gly753. The binding energy calculated for the T6P-TPP complex stood at −5.7 kcal/mol.

求助全文

通过发布文献求助，成功后即可免费获取论文全文。去求助

来源期刊

Protein expression and purification 生物-生化研究方法

CiteScore

3.70

自引率

6.20%

发文量

120

审稿时长

32 days

期刊介绍： Protein Expression and Purification is an international journal providing a forum for the dissemination of new information on protein expression, extraction, purification, characterization, and/or applications using conventional biochemical and/or modern molecular biological approaches and methods, which are of broad interest to the field. The journal does not typically publish repetitive examples of protein expression and purification involving standard, well-established, methods. However, exceptions might include studies on important and/or difficult to express and/or purify proteins and/or studies that include extensive protein characterization, which provide new, previously unpublished information.