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Reductase-catalyzed tetrahydrobiopterin regeneration alleviates the anti-competitive inhibition of tyrosine hydroxylation by 7,8-dihydrobiopterin
Catalysis Science & Technology ( IF 4.4 ) Pub Date : 2021-3-19 , DOI: 10.1039/d0cy01958e
Yinbiao Xu 1, 2, 3, 4, 5 , Youran Li 1, 2, 3, 4, 5 , Leyun Li 1, 2, 3, 4, 5 , Liang Zhang 1, 2, 3, 4, 5 , Zhongyang Ding 1, 2, 3, 4, 5 , Guiyang Shi 1, 2, 3, 4, 5
Catalysis Science & Technology ( IF 4.4 ) Pub Date : 2021-3-19 , DOI: 10.1039/d0cy01958e
Yinbiao Xu 1, 2, 3, 4, 5 , Youran Li 1, 2, 3, 4, 5 , Leyun Li 1, 2, 3, 4, 5 , Liang Zhang 1, 2, 3, 4, 5 , Zhongyang Ding 1, 2, 3, 4, 5 , Guiyang Shi 1, 2, 3, 4, 5
Affiliation
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L-Tyrosine hydroxylation by tyrosine hydroxylase is a significant reaction for preparing many nutraceutical and pharmaceutical chemicals. Two major challenges in constructing these pathways in bacteria are the improvement of hydroxylase catalytic efficiency and the production of cofactor tetrahydrobiopterin (BH4). In this study, we analyzed the evolutionary relationships and conserved protein sequences between tyrosine hydroxylases from different species by PhyML and MAFFT. Finally, we selected 7 tyrosine hydroxylases and 6 sepiapterin reductases. Subsequently, the function of different groups was identified by a combined whole-cell catalyst, and a series of novel tyrosine hydroxylase/sepiapterin reductase (TH/SPR) synthesis systems were screened including tyrosine hydroxylase (from Streptosporangium roseum DSM 43021 and Thermomonospora curvata DSM 43183) and sepiapterin reductase (from Photobacterium damselae, Chlorobaculum thiosulfatiphilum and Xenorhabdus poinarii), namely as SrTH/PdSPR, SrTH/CtSPR, SrTH/XpSPR and TcTH/PdSPR, which can synthesize L-Dopa by hydroxylating L-tyrosine in Bacillus licheniformis. Furthermore, we analyzed the characterization of SrTH by enzyme catalysis and demonstrated that 7,8-dihydrobiopterin (BH2) formed by BH4 autooxidation was an anticompetitive inhibitor on SrTH. Finally, pure dihydropteridine reductase from Escherichia coli (EcDHPR) was added to the solution, and L-Dopa could be continually synthesized after 3 h, which was improved by 86% at 6 h in the catalytic reaction by SrTH. This indicates that BH4 regeneration can alleviate the inhibition by BH2 during tyrosine hydroxylation. This study provides a good starting point and theoretical foundation for further modification to improve the catalytic efficiency of tyrosine hydroxylation by tyrosine hydroxylase.
中文翻译：
还原酶催化的四氢生物蝶呤再生减轻了7,8-二氢生物蝶呤对酪氨酸羟基化的反竞争抑制作用
酪氨酸羟化酶使L-酪氨酸羟化是制备许多保健食品和药物的重要反应。在细菌中构建这些途径的两个主要挑战是羟化酶催化效率的提高和辅因子四氢生物蝶呤（BH4）的产生。在这项研究中，我们通过PhyML和MAFFT分析了不同物种的酪氨酸羟化酶之间的进化关系和保守的蛋白质序列。最后，我们选择了7个酪氨酸羟化酶和6个sepaapterin还原酶。随后，通过组合的全细胞催化剂鉴定了不同基团的功能，并筛选了一系列新型的酪氨酸羟化酶/ septapterin还原酶（TH / SPR）合成系统，其中包括酪氨酸羟化酶（来自于Streptosporangium roseum）DSM 43021和Thermomonospora curvata DSM 43183）和sepiapterin还原酶（从发光菌damselae，Chlorobaculum thiosulfatiphilum和致病杆菌属poinarii），即作为SrTH / PdSPR，SrTH / CtSPR，SrTH / XpSPR和TcTH / PdSPR，其可以合成大号通过羟化-DOPA大号地衣芽孢杆菌中的酪氨酸。此外，我们通过酶催化分析了SrTH的特征，并证明了由BH4自氧化形成的7,8-二氢生物蝶呤（BH2）是对SrTH的抗竞争性抑制剂。最后，从纯二氢蝶啶还原酶大肠杆菌（EcDHPR）加入到该溶液中，和大号-Dopa可以在3 h后连续合成，在SrTH催化反应中6 h可以提高86％。这表明在酪氨酸羟基化过程中，BH4的再生可以减轻BH2的抑制作用。该研究为进一步改进酪氨酸羟化酶催化酪氨酸羟化效率提供了良好的起点和理论基础。
更新日期：2021-03-19
中文翻译：
还原酶催化的四氢生物蝶呤再生减轻了7,8-二氢生物蝶呤对酪氨酸羟基化的反竞争抑制作用
酪氨酸羟化酶使L-酪氨酸羟化是制备许多保健食品和药物的重要反应。在细菌中构建这些途径的两个主要挑战是羟化酶催化效率的提高和辅因子四氢生物蝶呤（BH4）的产生。在这项研究中，我们通过PhyML和MAFFT分析了不同物种的酪氨酸羟化酶之间的进化关系和保守的蛋白质序列。最后，我们选择了7个酪氨酸羟化酶和6个sepaapterin还原酶。随后，通过组合的全细胞催化剂鉴定了不同基团的功能，并筛选了一系列新型的酪氨酸羟化酶/ septapterin还原酶（TH / SPR）合成系统，其中包括酪氨酸羟化酶（来自于Streptosporangium roseum）DSM 43021和Thermomonospora curvata DSM 43183）和sepiapterin还原酶（从发光菌damselae，Chlorobaculum thiosulfatiphilum和致病杆菌属poinarii），即作为SrTH / PdSPR，SrTH / CtSPR，SrTH / XpSPR和TcTH / PdSPR，其可以合成大号通过羟化-DOPA大号地衣芽孢杆菌中的酪氨酸。此外，我们通过酶催化分析了SrTH的特征，并证明了由BH4自氧化形成的7,8-二氢生物蝶呤（BH2）是对SrTH的抗竞争性抑制剂。最后，从纯二氢蝶啶还原酶大肠杆菌（EcDHPR）加入到该溶液中，和大号-Dopa可以在3 h后连续合成，在SrTH催化反应中6 h可以提高86％。这表明在酪氨酸羟基化过程中，BH4的再生可以减轻BH2的抑制作用。该研究为进一步改进酪氨酸羟化酶催化酪氨酸羟化效率提供了良好的起点和理论基础。
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