Mechanism of Peptide Amidation: Structural and Kinetic Studies

肽酰胺化机制：结构和动力学研究

• 批准号: 0450465
• 负责人: L. Mario Amzel
• 金额: $ 71.95万
• 依托单位: Johns Hopkins University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-02-01 至 2010-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0450465&HistoricalAwards=false
• 关键词: Mechanism; Peptide; Amidation; Structural; Kinetic

Amidated peptides are fundamental signaling molecules found in species ranging from Aplysia to humans. Surprisingly, peptide amidation is not carried out by a transamination reaction, but by oxidative cleavage, glycine-extended precursors. A bifunctional enzyme, peptidylglycine alpha-amidating monooxygenase (PAM) catalyzes this reaction. The PAM gene encodes two domains (PHM and PAL) that catalyze two sequential reactions: alpha-hydroxylation of the glycine (PHM) and excision of the C alpha-N bond to give alpha-amidated peptide (PAL). PHM contains two redox active copper atoms that, upon reduction by ascorbate, catalyze the reduction of molecular oxygen for the hydroxylation of glycine-extended substrates. PAL is a zinc containing lyase that cleaves the C alpha-N bond after hydroxylation of the C alpha. This project will employ X-ray diffraction, kinetic experiments, peptide design, and site directed mutagenesis to elucidate important aspects of the mechanism of reactions catalyzed by PHM and PAL. Understanding the chemistry of PAM, especially that of the first step of the reaction, will not only contribute to the field of peptide amidation, but will also provide an outstanding paradigm for understanding long range electron transfer and the prevention of production of deleterious oxygen species. Broader Impacts: The PI of this project is actively involved in teaching as well as in bringing the excitement of science to underrepresented minorities. Mentoring of minority students will be achieved through participation in the Meyerhoff Scholars Programs of the University of Maryland Baltimore Campus. The project will also involve participation in the Science Day organized in conjunction with the Baltimore City school system.

胺化肽是从垂直到人类到人类的物种中发现的基本信号分子。 令人惊讶的是，肽胺不是通过透射反应而是通过氧化性裂解，甘氨酸延伸的前体进行的。 双功能酶，肽基甘氨酸α的单加氧酶（PAM）催化这种反应。 PAM基因编码两个催化两个顺序反应的域（PHM和PAL）：甘氨酸（PHM）的α-羟基化（PHM）和Cα-N键的切除以产生α增强的肽（PAL）。 PHM包含两个氧化还原活性铜原子，抗坏血酸还原后，催化了分子氧的还原，用于甘氨酸延伸的底物的羟基化。 PAL是一种含有裂解酶的锌，在Cα的羟基化后切割Cα-N键。 该项目将采用X射线衍射，动力学实验，肽设计和位点，定向诱变，以阐明PHM和PAL催化的反应机理的重要方面。了解PAM的化学，尤其是反应的第一步的化学性质，不仅会导致肽胺的领域，而且还将提供出色的范式，以理解远距离电子转移和预防有害氧气的产生。 更广泛的影响：该项目的PI积极参与教学以及将科学的兴奋带给代表性不足的少数群体。通过参加马里兰州巴尔的摩大学校园的Meyerhoff学者计划，将实现少数民族学生的指导。该项目还将涉及参加与巴尔的摩市学校系统一起组织的科学日。