A large number of enzymes expressed in both gram-positive and gram-negative bacteria are able to modify aminoglycoside antibiotics and confer resistance to their action. These enzymes have been divided based on their chemical group transformation. One such class is the acetyl transferases (AACs) - they catalyze the acetyl CoA-dependent acetylation of the amino group at the C3 position of the aminoglycoside.  This study focuses on a pair of acetyl transferases: The aminoglycoside-N3- acetyltransferase IIIb (AAC-IIIb) and the aminoglycoside-N3- acetyltransferase IIa (AAC-IIa).  AAC-IIa and AAC-IIIb are very similar in their amino acid sequence and structure and yet have a strong difference in their substrate selectivity, kinetic and thermodynamic properties. Nuclear Magnetic Resonance (NMR) data suggests that when AAC-IIa is in its apo form it has a very dynamic nature, seen by a number of overlapping peaks whereas the AAC-IIIb shows a well disperse spectrum and it is much less dynamic. AAC-IIa adopts a well-defined structure in solution only with bound antibiotics as detected by NMR.   Analytical ultracentrifugation (AUC) studies showed a major difference between these two enzymes; while AAC-IIa is a monomer, AAC-IIIb is a dimer in solution.           The differential scanning calorimetry (DSC) studies showed that, unlike AAC-IIIb, binding of aminoglycosides to AAC-IIa didn't alter the Tm. Studies using the isothermal titration calorimetry (ITC) showed that binding of two structurally very similar aminoglycosides, neomycin and paromomycin, to AAC-IIIb, caused opposing effects in heat capacity change and radius of hydration of the complex. Temperature dependence of the binding of aminoglycosides to AAC-IIa revealed that, unlike AAC-IIIb, above 22�C, the binding enthalpy includes strong contributions from folding enthalpy.  Below 22�C, the binding of aminoglycosides to this enzyme shows a heat capacity change consistent with carbohydrate-protein interactions and the binding enthalpy does not appear have contribution of the folding enthalpy.

Strikingly different however alike: The two aminoglycoside acetyl transferases

248th National Meeting (2014)

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.