Ligand promiscuity through the eyes of the aminoglycoside N3 acetyltransferase IIa

Authors

  • Adrianne L. Norris,

    1. From the Department of Biochemistry and Cellular and Molecular Biology, The University of Tennessee, Knoxville, Tennessee
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  • Engin H. Serpersu

    Corresponding author
    1. Graduate School of Genome Science and Technology, University of Tennessee and Oak Ridge National Laboratories, Knoxville, Tennessee
    • From the Department of Biochemistry and Cellular and Molecular Biology, The University of Tennessee, Knoxville, Tennessee
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  • All work presented herein was performed at the University of Tennessee, Knoxville.

Correspondence to: Engin H. Serpersu, Department of Biochemistry Cell and Molecular Biology, The University of Tennessee, M407 Walters Life Sciences Bldg., Knoxville, TN 37996-0840. E-mail: Serpersu@utk.edu

Abstract

Aminoglycoside-modifying enzymes (AGMEs) are expressed in many pathogenic bacteria and cause resistance to aminoglycoside (AG) antibiotics. Remarkably, the substrate promiscuity of AGMEs is quite variable. The molecular basis for such ligand promiscuity is largely unknown as there is not an obvious link between amino acid sequence or structure and the antibiotic profiles of AGMEs. To address this issue, this article presents the first kinetic and thermodynamic characterization of one of the least promiscuous AGMEs, the AG N3 acetyltransferase-IIa (AAC-IIa) and its comparison to two highly promiscuous AGMEs, the AG N3-acetyltransferase-IIIb (AAC-IIIb) and the AG phosphotransferase(3′)-IIIa (APH). Despite having similar antibiotic selectivities, AAC-IIIb and APH catalyze different reactions and share no homology to one another. AAC-IIa and AAC-IIIb catalyze the same reaction and are very similar in both amino acid sequence and structure. However, they demonstrate strong differences in their substrate profiles and kinetic and thermodynamic properties. AAC-IIa and APH are also polar opposites in terms of ligand promiscuity but share no sequence or apparent structural homology. However, they both are highly dynamic and may even contain disordered segments and both adopt well-defined conformations when AGs are bound. Contrary to this AAC-IIIb maintains a well-defined structure even in apo form. Data presented herein suggest that the antibiotic promiscuity of AGMEs may be determined neither by the flexibility of the protein nor the size of the active site cavity alone but strongly modulated or controlled by the effects of the cosubstrate on the dynamic and thermodynamic properties of the enzyme.

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