Molecular Basis Of Ligand Selectivity In Aminoglycoside Acetyltransferases

Resistant bacteria employ drug modifying enzymes like aminoglycoside acetyltransferases (AACs) to evade the effects of aminoglycoside antibiotics. Even though the AACs share a high level of sequence and structural similarity, they exhibit a range of substrate profiles. Thus, these enzymes present an excellent opportunity to study the molecular principles underlying substrate promiscuity and selectivity. AAC-VIa and AAC-IIIb possess the same structural fold, catalyze the same modification reaction and utilize the same catalytic machinery to do so. Despite these similarities, they exhibit quite different substrate profiles. AAC-VIa, is a highly selective AAC, capable of modifying only five aminoglycosides, limited to kanamycin class of drugs. AAC-IIIb, on the other hand, is a highly promiscuous acetyltransferase and can modify upto 15 different aminoglycosides, from both the neomycin and kanamycin classes. Structure based sequence alignment shows that AAC-VIa has a loop with a four-residue insertion, which is absent in AAC-IIIb. Instead, this region in AAC-IIIb is occupied by a subsite of the binding site, utilized to bind neomycin class of aminoglycosides, which are not catalyzed by AAC-VIa. Unlike AAC-IIIb, AAC-VIa is able to kinetically distinguish between its substrates. This is accomplished by more preferable ligands binding in a different conformation compared to the lesser preferable ligands. AAC-IIIb utilizes a larger and bipartite binding site, to allow the binding of structurally varied substrates and catalyzes all of them with similar efficiency. These structures provide structural framework for the unusual thermodynamic properties of enzyme-ligand complexes of AAC-IIIb. Overall, these studies shed light on molecular principles underlying ligand selection by AACs and would contribute towards the design and development of more effective antibacterial drugs.