通过基于代和突变发生的定向进化来操纵烯环氧化酶的立体选择性
Huabao Zheng1, Manfred T Reetz
1Max-Planck-Institut für Kohlenforschung, Kaiser-Wilhelm-Platz 1, 45470 Mülheim an der Ruhr, Germany.
Journal of the American Chemical Society
|October 21, 2010
概括
工程化环氧化酶 (EH) 酶具有广泛的基质范围和高立体选择性,用于不对称的合成. 代和突变性迅速为各种环氧化物转换创造了新的EH变体.
科学领域:
- 生物催化剂是一种生物催化剂.
- 酵素工程是什么意思 酵素工程
- 有机合成 有机合成
背景情况:
- 来自Rhodococcus erythropolis DCL 14的林烯环氧化酶 (LEH) 具有独特的结构和机械性质.
- 由于LEH的天然基质特异性,限制了其在其他环氧化物不对称合成中的应用.
- 存在对具有更广泛基质范围和高立体选择性的工程环氧化酶的需求.
研究的目的:
- 通过使用定向进化来设计具有增强基质范围和立体选择性的LEH突变体.
- 为生物催化剂开发开发开发一种快速可靠的蛋白质工程方法.
主要方法:
- 代和突变发生法 (ISM) 用于修改LEH结合口袋.
- 结合口袋中的残留物被分组并使用减少的氨基酸字母表进行突变发生.
- 使用定向进化来选改进的LEH变体.
主要成果:
- 工程LEH突变体证明了对 (R,R) -或 (S,S) -二醇具有高立体选择性的环二氧化物去对称的能力.
- 进化突变物有效催化了其他氧化物脱对称和种族基质的动态分辨率.
- 通过对不到5000个变压器进行选,取得了成功的工程.
结论:
- 代和突变发生是一种快速有效的方法,用于环氧化酸酶的蛋白质工程.
- 工程LEH变体表现出广泛的基质范围和高立体选择性,扩大了它们在不对称合成中的实用性.
- 该研究验证了ISM用于开发用于各种化学转换的生物催化剂.
相关概念视频
02:57Sharpless Epoxidation
The conversion of allylic alcohols into epoxides using the chiral catalyst was discovered by K. Barry Sharpless and is known as Sharpless epoxidation. The use of a chiral catalyst enables the formation of one enantiomer of the product in excess. This chiral catalyst is mainly a chiral complex of titanium tetraisopropoxide and tartrate ester (specific stereoisomer). The stereoisomer used in the chiral catalyst dictates the formation of the enantiomer of the product. In other words, the use of...
01:22Bioreactor Controls-III
Strain improvement is a foundational strategy in industrial microbiology aimed at maximizing microbial productivity, particularly because natural isolates typically yield commercially valuable products in very low concentrations. Although optimizing the culture medium and environmental conditions can improve yields, these adjustments are inherently limited by the organism’s genetic potential. As a result, the focus shifts toward genetic modifications to enhance biosynthetic capacity. The...
01:24Evolution of New Traits in Microbes
Microorganisms evolve rapidly due to their large population sizes and short generation times, often exhibiting measurable changes within days under laboratory conditions. Natural selection acts on standing genetic variation, enabling the retention and amplification of beneficial traits that confer fitness advantages in changing environments.Adaptive Pigment Regulation in RhodobacterIn Rhodobacter, a genus of purple non-sulfur bacteria, light-harvesting pigments such as bacteriochlorophyll and...
02:24Acid-Catalyzed Ring-Opening of Epoxides
Epoxides that are three-membered ring systems are more reactive than other cyclic and acyclic ethers. The high reactivity of epoxides originates from the strain present in the ring. This ring strain acts as a driving force for epoxides to undergo ring-opening reactions either with halogen acids or weak nucleophiles in the presence of mild acid. The acid catalyst converts the epoxide oxygen, a poor leaving group, into an oxonium ion, a better leaving group, making the reaction feasible. The...
02:34Regioselectivity and Stereochemistry of Acid-Catalyzed Hydration
The rate of acid-catalyzed hydration of alkenes depends on the alkene's structure, as the presence of alkyl substituents at the double bond can significantly influence the rate.
03:00Preparation of Epoxides
Overview
Epoxides result from alkene oxidation, which can be achieved by a) air, b) peroxy acids, c) hypochlorous acids, and d) halohydrin cyclization.
Epoxidation with Peroxy Acids
Epoxidation of alkenes via oxidation with peroxy acids involves the conversion of a carbon–carbon double bond to an epoxide using the oxidizing agent meta-chloroperoxybenzoic acid, commonly known as MCPBA. Since the O–O bond of peroxy acids is very weak, the addition of electrophilic oxygen of peroxy acids to...
Epoxides result from alkene oxidation, which can be achieved by a) air, b) peroxy acids, c) hypochlorous acids, and d) halohydrin cyclization.
Epoxidation with Peroxy Acids
Epoxidation of alkenes via oxidation with peroxy acids involves the conversion of a carbon–carbon double bond to an epoxide using the oxidizing agent meta-chloroperoxybenzoic acid, commonly known as MCPBA. Since the O–O bond of peroxy acids is very weak, the addition of electrophilic oxygen of peroxy acids to...