Structure-function analyses on AcuB encoded by the Bacillus subtilis acuABC-operon involved in regulation of acetyl-CoA synthetase by post-translational lysine acetylation

枯草芽孢杆菌 acuABC-操纵子编码的 AcuB 的结构功能分析参与翻译后赖氨酸乙酰化调节乙酰辅酶 A 合成酶

• 批准号: 534243417
• 负责人: Professor Dr. Michael Lammers
• 金额: --
• 依托单位: Institut für Biochemie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/534243417?language=en
• 关键词: Structure; function; analyses; AcuB; encoded

Bacterial cells have to cope with rapidly changing environmental conditions, including alterations in nutrient availability. A way to sense the cellular metabolic state is the regulation of protein function by post-translational lysine acetylation. In bacteria, lysine acetylation is enzymatically catalyzed by lysine acetyltransferases (KATs) that use acetyl-CoA as donor molecule for the acetylation of lysine side chains. Acetylated lysine side chains can be deacetylated by deacetylases, which are classified into two groups, i.e. the classical Zn2+-dependent deacetylases (KDACs) and the NAD+-dependent sirtuins. Next to its role as donor molecule for acetylation, acetyl-CoA is a central metabolite. Acetyl-CoA can be, amongst other, generated by the activity of AMP-forming acetyl-CoA synthetases (Acs). This reaction is split into two half reactions. In the 1st half reaction, the adenylation reaction, acetate is activated by adenylation to yield acetyl-AMP. Subsequently, in the 2nd half reaction, acetyl-AMP is converted to the thioester acetyl-CoA (thioester formation). In the Gram-positive bacterial species Bacillus subtilis, the acetyl-CoA synthetase AcsA is encoded upstream from the acuABC-operon and is reversely transcribed. The acuABC-operon was originally identified to be involved in butanediol and acetoin catabolism. The function of the gene products of the acuABC operon are mechanistically only partially understood. The gene acuA encodes for a GNAT-type KAT to acetylate AcsA at a C-terminal lysine side chain, i.e. K549, and thereby inactivate AcsA activity. AcuC constitutes a classical Zn2+-dependent deacetylase. It is known that AcuC is capable to deacetylate and thereby (re-)activate AcsA. The exact molecular mechanisms underlying the inactivation of AcsA activity by AcuA mediated K549-acetylation are only marginally understood. The role of AcuB in the interplay of AcuA, AcuC and AcsA is totally unclear. Interestingly, other bacterial species such as Staphylococcus aureus share a similar gene organization. However, in S. aureus the operon encodes only for AcuA and AcuC but the acuB gene is missing. This suggests that AcuB plays a role that might be specific for some bacterial strains. We postulate that AcuB, which is completely functionally uncharacterized, might be an important regulator for this interplay, i.e. sensing the metabolic state, adjusting AcuA and/or AcuC activity to modulate AcsA activity. In B. subtilis, AcuB by binding to AcuC and/or AcuA and to molecules such as NAD+, AMP, ATP or CoA/acetyl-CoA might be the regulator that adapts the activity of the deactylase AcuC and/or the acetyltransferase AcuA to the cellular metabolic state enabling to coordinate AcsA activity. In this project we want to focus on the exact functional role of AcuB that is encoded by the acuABC-operon in B. subtilis. These data will reveal if regulation of classical deacetylases by binding to sensory proteins is a more general regulatory system.

细菌细胞必须应对迅速变化的环境条件，包括养分可用性的改变。感知细胞代谢态的一种方法是通过翻译后赖氨酸乙酰化来调节蛋白质功能。在细菌中，赖氨酸乙酰化是由赖氨酸乙酰基转移酶（KAT）酶催化的，该赖氨酸使用乙酰辅酶A作为供体分子作为赖氨酸侧链的乙酰化。乙酰化的赖氨酸侧链可以通过脱乙酰基酶脱乙酰基，分为两组，即经典的Zn2+依赖性脱乙酰基酶（KDACS）和NAD+依赖性sirtuins。在其作为乙酰化供体分子的作用之后，乙酰辅酶A是一种中央代谢物。除其他外，乙酰辅酶A可以由AMP形成的乙酰-COA合成酶（AC）的活性产生。该反应分为两个一半反应。在第一个半反应中，腺苷酸化反应，乙酸乙酸酯通过腺苷酸化激活以产生乙酰基AMP。随后，在第二半反应中，乙酰基AMP转化为硫酯乙酰辅酶A（硫酯形成）。在革兰氏阳性的细菌种类杆菌中，乙酰辅酶A合成酶ACSA是从Acuabc-Operon上游编码的，并反复转录。 Acuabc-Operon最初被确定为与丁二醇和乙酸分解代谢有关。 Acuabc操纵子的基因产物的功能在机械上仅部分理解。基因ACUA在C末端赖氨酸侧链（即K549）中编码GNAT型KAT对乙酰酸ACSA进行编码，从而使ACSA活性失活。 ACUC构成经典的Zn2+依赖性脱乙酰基酶。众所周知，ACUC能够脱乙酰酸，从而（重新）激活ACSA。 ACUA介导的K549-乙酰化对ACSA活性失活的确切分子机制仅略得多。 ACUB在ACUA，ACUC和ACSA相互作用中的作用尚不清楚。有趣的是，金黄色葡萄球菌等其他细菌物种具有类似的基因组织。然而，在金黄色葡萄球菌中，操纵子仅编码为ACUA和ACUC，但缺少ACUB基因。这表明ACUB扮演的作用可能是某些细菌菌株的特定作用。我们假设ACUB在功能上完全没有表征，可能是该相互作用的重要调节剂，即感测代谢态，调整ACUA和/或ACUC活性以调节ACSA活性。在枯草芽孢杆菌中，ACUB与ACUC和/或ACUA结合，以及与NAD+，AMP，ATP或COA/COA/乙酰-COA等分子，可能是适应脱链基酶ACUC的活性和/或乙酰基转移酶ACUA的活性，对细胞代谢呈辅助ACSA的活性。在这个项目中，我们希望关注由枯草芽孢杆菌中Acuabc-Operon编码的ACUB的确切功能作用。这些数据将揭示是否通过与感觉蛋白结合对经典脱乙酰基酶的调节是更一般的调节系统。