Insight into the 3D structure and substrate specificity of previously uncharacterized GNAT superfamily acetyltransferases from pathogenic bacteria.

Karolina A Majorek, Tomasz Osinski, David T Tran, Alina Revilla, Wayne F Anderson, Wladek Minor, Misty L Kuhn

Biochimica et biophysica acta. Proteins and proteomics 2017 Jan

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Members of the Gcn5-related N-acetyltransferase (GNAT) superfamily catalyze the acetylation of a wide range of small molecule and protein substrates. Due to their abundance in all kingdoms of life and diversity of their functions, they are implicated in many aspects of eukaryotic and prokaryotic physiology. Although numerous GNATs have been identified thus far, many remain structurally and functionally uncharacterized. The elucidation of their structures and functions is critical for broadening our knowledge of this diverse and important superfamily. In this work, we present the structural and kinetic analyses of two previously uncharacterized bacterial acetyltransferases - SACOL1063 from Staphylococcus aureus strain COL and CD1211 from Clostridium difficile strain 630. Our structures of SACOL1063 show substantial flexibility of a loop that is likely responsible for substrate recognition and binding compared to structures of other homologs. In the CoA complex structure, we found two CoA molecules bound in both the canonical AcCoA/CoA-binding site and the acceptor-substrate-binding site. Our work also provides initial clues regarding the substrate specificity of these two enzymes; however, their native function(s) remain unknown. We found both proteins act as N- rather than O-acetyltransferases and preferentially acetylate l-threonine. The combination of structural and kinetic analyses of these two previously uncharacterized GNATs provides fundamental knowledge and a framework on which future studies can be built to elucidate their native functions. Copyright Â© 2016 Elsevier B.V. All rights reserved.

Citation

Karolina A Majorek, Tomasz Osinski, David T Tran, Alina Revilla, Wayne F Anderson, Wladek Minor, Misty L Kuhn. Insight into the 3D structure and substrate specificity of previously uncharacterized GNAT superfamily acetyltransferases from pathogenic bacteria. Biochimica et biophysica acta. Proteins and proteomics. 2017 Jan;1865(1):55-64