Structure and Function of Hexapeptide Acyltransferases

六肽酰基转移酶的结构和功能

• 批准号: 6999799
• 负责人: STEVEN L RODERICK
• 金额: $ 28.28万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-12-01 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6999799
• 关键词: Enterococcus; active sites; acyltransferase; antibacterial agents; bacterial proteins; crystallization; enzyme inhibitors; enzyme mechanism; enzyme substrate complex; high throughput technology; ligands; microorganism culture; mutant; protein purification; protein structure function; site directed mutagenesis; thermodynamics

DESCRIPTION (provided by applicant): Hexapeptide acyltransferases are characterized by tandem repeated copies of a six residue 'hexapeptide repeat' periodicity theme that encode folding of a unique left-handed parallel beta-helix structural domain. These enzymes play important roles in detoxification, cell wall formation and amino acid biosynthesis, but are absent in humans. Three bacterial hexapeptide acetyltransferases play well understood biological roles, but have not been mechanistically characterized or developed as antimicrobial drug targets. The gram-positive streptogramin acetyltransferase (VatD) inactivates the group A component of streptogramin mixtures currently used as last resort antibiotics in human medicine. High throughput screening and structure determination of enzyme-ligand complexes will be used to characterize the interaction of inhibitors with Vat(D). These compounds, or their synthetic variants, may find use as antibiotics that restore the susceptibility of gram positive pathogens to existing streptogramin mixtures. The bifunctional uridyltransferase (GImU) is responsible for the biosynthesis of UDP-GlcNAc, an essential building block for both the peptidoglycan and lipopolysaccharide components of bacterial cell walls. Its acetyltransferase active site is unique but has not been mechanistically characterized. Its structure and mechanism of action will be defined in order to develop the potential of GlmU as an attractive mechanism-based antibacterial target. Serine transacetylase (STA) O-acetylates serine in the cysteine biosynthetic pathway of bacteria and is the key regulatory enzyme of cysteine biosynthesis and sulfate assimilation. The structure and reaction mechanism of STA will be studied as a resolved enzyme and as a component of the binary cysteine synthase complex. STA is inhibited by cysteine, but must also utilize serine. The means by which STA recognizes these isosteric amino acids as inhibitor and substrate, respectively, may point to features of antimicrobial agents that duplicate this natural high-affinity mode of inhibition. The successful completion of these aims will serve to advance basic functional understanding of the hexapeptide acetyltransferase superfamily of enzymes and will develop their application as screening-based or mechanism-based targets for antibacterial drug design.

描述（由申请人提供）：六肽酰基转移酶的特征在于六残基“六肽重复”周期性主题的串联重复拷贝，其编码独特的左手平行β-螺旋结构域的折叠。这些酶在解毒、细胞壁形成和氨基酸生物合成中发挥重要作用，但在人体中不存在。三种细菌六肽乙酰转移酶发挥着众所周知的生物学作用，但尚未被机械表征或开发为抗菌药物靶点。 革兰氏阳性链阳菌素乙酰转移酶 (VatD) 可使链阳菌素混合物中的 A 组成分失活，链阳菌素混合物目前用作人类医学中最后的抗生素。酶-配体复合物的高通量筛选和结构测定将用于表征抑制剂与 Vat(D) 的相互作用。这些化合物或其合成变体可以用作抗生素，恢复革兰氏阳性病原体对现有链阳菌素混合物的敏感性。 双功能尿苷基转移酶 (GImU) 负责 UDP-GlcNAc 的生物合成，UDP-GlcNAc 是细菌细胞壁肽聚糖和脂多糖成分的重要组成部分。其乙酰转移酶活性位点是独特的，但尚未对其进行机械表征。其结构和作用机制将被定义，以开发 GlmU 作为有吸引力的基于机制的抗菌靶点的潜力。 丝氨酸转乙酰酶（STA）O-乙酰化细菌半胱氨酸生物合成途径中的丝氨酸，是半胱氨酸生物合成和硫酸盐同化的关键调节酶。 STA 的结构和反应机制将作为解析酶和二元半胱氨酸合酶复合物的组成部分进行研究。 STA 被半胱氨酸抑制，但也必须利用丝氨酸。 STA 将这些等排氨基酸分别识别为抑制剂和底物的方法可能表明抗菌剂的特征复制了这种天然的高亲和力抑制模式。 这些目标的成功完成将有助于增进对六肽乙酰转移酶超家族的基本功能理解，并将开发其作为基于筛选或基于机制的抗菌药物设计靶标的应用。