Structural and Mechanistic Analysis of Drosophila melanogaster Agmatine N-Acetyltransferase, an Enzyme that Catalyzes the Formation of N-Acetylagmatine

Autor: Daniel R. Dempsey, Derek A. Nichols, Matthew R. Battistini, Orville Pemberton, Santiago Rodriguez Ospina, Xiujun Zhang, Anne-Marie Carpenter, Brian G. O’Flynn, James W. Leahy, Ankush Kanwar, Eric M. Lewandowski, Yu Chen, David J. Merkler
Jazyk: angličtina
Rok vydání: 2017
Předmět:
Zdroj: Scientific Reports, Vol 7, Iss 1, Pp 1-15 (2017)
Druh dokumentu: article
ISSN: 2045-2322
DOI: 10.1038/s41598-017-13669-6
Popis: Abstract Agmatine N-acetyltransferase (AgmNAT) catalyzes the formation of N-acetylagmatine from acetyl-CoA and agmatine. Herein, we provide evidence that Drosophila melanogaster AgmNAT (CG15766) catalyzes the formation of N-acetylagmatine using an ordered sequential mechanism; acetyl-CoA binds prior to agmatine to generate an AgmNAT•acetyl-CoA•agmatine ternary complex prior to catalysis. Additionally, we solved a crystal structure for the apo form of AgmNAT with an atomic resolution of 2.3 Å, which points towards specific amino acids that may function in catalysis or active site formation. Using the crystal structure, primary sequence alignment, pH-activity profiles, and site-directed mutagenesis, we evaluated a series of active site amino acids in order to assign their functional roles in AgmNAT. More specifically, pH-activity profiles identified at least one catalytically important, ionizable group with an apparent pKa of ~7.5, which corresponds to the general base in catalysis, Glu-34. Moreover, these data led to a proposed chemical mechanism, which is consistent with the structure and our biochemical analysis of AgmNAT.
Databáze: Directory of Open Access Journals
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