基质捕获在多基酸合成酶和酸酶域中:巨乳素形成的结构基础
Tyler M McCullough1,2, Vishakha Choudhary1,2, David L Akey1
1Life Sciences Institute, Mary Sue Coleman Hall, 210 Washtenaw Ave., University of Michigan, Ann Arbor, MI 48109-2216, United States.
概括
研究人员使用1,3-二氨基酸 (DAP) 来捕获中间体,设计了类型二化酶 (TEs),揭示了这些酶如何形成巨乳抗生素. 这项工作有助于开发用于药物发现的新生物催化剂.
科学领域:
- 生物化学 生物化学
- 分子生物学分子生物学
- 合成生物学 合成生物学
背景情况:
- 抗生素耐药性需要开发新的抗菌药物,特别是宏类药物.
- 聚基化合成酶 (PKS) 化酶 (TE) 在化物生物合成中对宏乐拉克支架形成至关重要.
- 对 TE 机制的理解有限,这阻碍了它们作为多功能生物催化剂的使用.
研究的目的:
- 阐明TE选择性在巨乳素形成中的机制.
- 设计TE以适应各种自然和非自然基质.
- 为 TE 工程和优化提供见解.
主要方法:
- 通过用1,3-diaminopropionic acid (DAP) 替换活性位点氨酸氧化物,将乙烯酸酶中间体作为稳定胺基被捕获.
- 从皮克罗米辛和红色素途径生成和净化DAP修饰的TE (DAPTE).
- 测试了TEDAP变体与各种多基基板,并确定了晶体结构.
主要成果:
- 红色素TE表现出允许的基质选择性,而皮克罗米辛TE对其本土基质具有选择性.
- 皮克罗米辛TEDAP基质复合物的晶体结构揭示了一个卷曲的heptaketide,与TE的乙烯腔高度互补.
- 在不同的TEs中观察到明显的乙腔形状,包括尤文尼米辛,泰洛辛和流血素通路.
结论:
- TEs通过在它们的乙腔内的特定基质相互作用来控制巨乳素的形成.
- 使用像DAP这样非自然氨基酸的工程TE提供了机械洞察力和生物催化剂开发的潜力.
- TE 乙腔的结构多样性为设计具有针对抗生素合成的基质特异性的 TE 提供了基础.
相关概念视频
Regioselective Formation of Enolates
2.5K
As depicted in the figure below, the unsymmetrical ketones can form two possible enolates: less substituted or more substituted enolates. Usually, the thermodynamic enolates are formed from the more substituted α-carbon atom, while the kinetic enolates are formed faster by deprotonation from the less substituted position. The thermodynamic enolates have lower energy, so they are more stable. But the energy required to form kinetic enolates is less.
2.5K
Keto–Enol Tautomerism: Mechanism
5.1K
The keto and enol forms are known as tautomers and they constantly interconvert (or tautomerize) between the two forms under acid or base catalyzed conditions. Both the reactions involve the same steps—protonation and deprotonation— although in the reverse order.
5.1K
Feedback Inhibition
53.6K
Biochemical reactions are occurring constantly in cells, converting starting substances to different products, usually with the help of enzymes that speed the reactions. Without enzymes, it would take far too long for most reactions to occur to be useful to the cell!
53.6K
Intramolecular Claisen Condensation of Dicarboxylic Esters: Dieckmann Cyclization
2.3K
Dieckmann cyclization is an intramolecular Claisen condensation of diesters. The reaction occurs in the presence of a base and generates a cyclic β-ketoester as the final product. Commonly, 1, 6 and 1, 7-diesters are preferred substrates for the reaction since the generated five, and six-membered cyclic β-keto esters are particularly more stable.
2.3K
Esters to β-Ketoesters: Claisen Condensation Mechanism
3.5K
Regular Claisen condensation involves the synthesis of β-ketoesters by combining identical ester molecules bearing two α hydrogens in the presence of an alkoxide base. The reaction commences with the deprotonation of the acidic α hydrogen by the base to form a resonance stabilized ester enolate. This nucleophilic ion then attacks the carbonyl center of another ester molecule to generate a tetrahedral alkoxide intermediate. Next, the expulsion of the alkoxide group from the...
3.5K
Alkylation of β-Diester Enolates: Malonic Ester Synthesis
3.3K
Malonic ester synthesis is a method to obtain α substituted carboxylic acids from ꞵ-diesters such as diethyl malonate and alkyl halides.
3.3K


