Domain-domain interactions in the aminoglycoside antibiotic resistance enzyme AAC(6')-APH(2'').
The most common determinant of aminoglycoside antibiotic resistance in Gram positive bacterial pathogens, such as Staphylococcus aureus, is a modifying enzyme, AAC(6')-APH(2' '), capable of acetylating and phosphorylating a wide range of antibiotics. This enzyme is unique in that it is composed of two separable modification domains, and although a number of studies have been conducted on the acetyltransferase and phosphotransferase activities in isolation, little is known about the role and impact of domain interactions on antibiotic resistance. Kinetic analysis and in vivo assessment of a number of N- and C-terminal truncated proteins have demonstrated that the two domains operate independently and do not accentuate one another's resistance activity. However, the two domains are structurally integrated, and mutational analysis has demonstrated that a predicted connecting alpha-helix is especially critical for maintaining proper structure and function of both activities. AAC(6')-APH(2' ') detoxifies a staggering array of aminoglycosides, where one or both activities make important contributions depending on the antibiotic. Thus, to overcome antibiotic resistance associated with AAC(6')-APH(2' '), aminoglycosides resistant to modification and/or inhibitors against both activities must be employed. Domain-domain interactions in AAC(6')-APH(2' ') offer a unique target for inhibitor strategies, as we show that their disruption simultaneously inhibits both activities >90%.
Boehr
David D
DD
Antimicrobial Research Centre, Department of Biochemistry, McMaster University, 1200 Main Street West, Hamilton, Ontario, Canada, L8N 3Z5.
Daigle
Denis M
DM
Wright
Gerard D
GD
D000123:Acetyltransferases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D000617:Aminoglycosides / Q000037:antagonists & inhibitors / Q000737:chemistry*; D000900:Anti-Bacterial Agents / Q000037:antagonists & inhibitors / Q000737:chemistry*; D001412:Bacillus subtilis / Q000201:enzymology; D015153:Blotting, Western; D002850:Chromatography, Gel; D004252:DNA Mutational Analysis; D024881:Drug Resistance, Bacterial / Q000235:genetics; D013293:Enterococcus faecalis / Q000201:enzymology; D004795:Enzyme Stability / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology; D006160:Guanosine Triphosphate / Q000737:chemistry; D006358:Hot Temperature; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D009097:Multienzyme Complexes / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D010446:Peptide Fragments / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D017433:Protein Structure, Secondary / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D017384:Sequence Deletion; D013211:Staphylococcus aureus / Q000201:enzymology
D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D009097:Multienzyme Complexes; D010446:Peptide Fragments; D006160:Guanosine Triphosphate; C108453:6'-aminoglycoside acetyltransferase-2''-aminoglycoside phosphotransferase; D000123:Acetyltransferases; D017853:Phosphotransferases (Alcohol Group Acceptor)