title: Structural and Biochemical Characterization of Acinetobacter spp. Aminoglycoside Acetyltransferases Highlights Functional and Evolutionary Variation among Antibiotic Resistance Enzymes.
abstract: Modification of aminoglycosides by N-acetyltransferases (AACs) is one of the major mechanisms of resistance to these antibiotics in human bacterial pathogens. More than 50 enzymes belonging to the AAC(6') subfamily have been identified in Gram-negative and Gram-positive clinical isolates. Our understanding of the molecular function and evolutionary origin of these resistance enzymes remains incomplete. Here we report the structural and enzymatic characterization of AAC(6')-Ig and AAC(6')-Ih from Acinetobacter spp. The crystal structure of AAC(6')-Ig in complex with tobramycin revealed a large substrate-binding cleft remaining partially unoccupied by the substrate, which is in stark contrast with the previously characterized AAC(6')-Ib enzyme. Enzymatic analysis indicated that AAC(6')-Ig and -Ih possess a broad specificity against aminoglycosides but with significantly lower turnover rates as compared to other AAC(6') enzymes. Structure- and function-informed phylogenetic analysis of AAC(6') enzymes led to identification of at least three distinct subfamilies varying in oligomeric state, active site composition, and drug recognition mode. Our data support the concept of AAC(6') functionality originating through convergent evolution from diverse Gcn5-related-N-acetyltransferase (GNAT) ancestral enzymes, with AAC(6')-Ig and -Ih representing enzymes that may still retain ancestral nonresistance functions in the cell as provided by their particular active site properties.
authors: Department of Chemical Engineering and Applied Chemistry, University of Toronto , 200 College Street, Toronto, Ontario M5G 1L6, Canada.
authors: Peter J
authors: PJ
authors: Stogios
authors: Department of Chemistry and Biochemistry, San Francisco State University , San Francisco, California 94132, United States.
authors: Misty L
authors: ML
authors: Kuhn
authors: Department of Chemical Engineering and Applied Chemistry, University of Toronto , 200 College Street, Toronto, Ontario M5G 1L6, Canada.
authors: Elena
authors: E
authors: Evdokimova
authors: Department of Chemistry and Biochemistry, San Francisco State University , San Francisco, California 94132, United States.
authors: Melissa
authors: M
authors: Law
authors: Institut Pasteur , Unité des Agents Antibactériens, 25 rue du Docteur Roux, 75724 Cedex 15 Paris, France.
authors: Patrice
authors: P
authors: Courvalin
authors: Department of Chemical Engineering and Applied Chemistry, University of Toronto , 200 College Street, Toronto, Ontario M5G 1L6, Canada.
authors: Alexei
authors: A
authors: Savchenko
mesh_terms: D000123:Acetyltransferases / Q000737:chemistry* / Q000378:metabolism*; D000150:Acinetobacter / Q000737:chemistry / Q000201:enzymology*; D000617:Aminoglycosides / Q000494:pharmacology; D018360:Crystallography, X-Ray; D004352:Drug Resistance, Microbial; D019143:Evolution, Molecular; D008958:Models, Molecular; D010802:Phylogeny; D011485:Protein Binding; D011487:Protein Conformation; D014031:Tobramycin / Q000494:pharmacology
chemical_list: D000617:Aminoglycosides; D000123:Acetyltransferases; C019681:aminoglycoside acetyltransferase; D014031:Tobramycin
keywords: Acinetobacter; Gcn5-related N-acetyltransferase; aminoglycoside; antibiotic resistance; crystal structure