Please use this identifier to cite or link to this item: https://doi.org/10.1038/srep37061
Title: Ligand-mediated changes in conformational dynamics of NpmA: Implications for ribosomal interactions
Authors: NILOFER HUSAIN 
Tulsian N.K. 
Chien W.L. 
Suresh S. 
Anand G.S.
Sivaraman J. 
Keywords: Escherichia coli protein
ligand
methyltransferase
NpmA protein, E coli
s adenosylhomocysteine
s adenosylmethionine
bacterial 30S ribosomal subunit
binding site
chemistry
deuterium hydrogen exchange
enzymology
Escherichia coli
mass spectrometry
metabolism
Binding Sites
Deuterium Exchange Measurement
Escherichia coli
Escherichia coli Proteins
Ligands
Mass Spectrometry
Methyltransferases
Ribosome Subunits, Small, Bacterial
S-Adenosylhomocysteine
S-Adenosylmethionine
Issue Date: 2016
Citation: NILOFER HUSAIN, Tulsian N.K., Chien W.L., Suresh S., Anand G.S., Sivaraman J. (2016). Ligand-mediated changes in conformational dynamics of NpmA: Implications for ribosomal interactions. Scientific Reports 6 : 37061. ScholarBank@NUS Repository. https://doi.org/10.1038/srep37061
Abstract: Aminoglycosides are broad-spectrum antibiotics that bind to the 30S ribosomal subunit (30S) of bacteria and disrupt protein translation. NpmA, a structurally well-characterized methyltransferase identified in an E. coli clinical isolate, catalyzes methylation of 30S at A1408 of the 16S rRNA and confers aminoglycoside resistance. Using sucrose cushion centrifugation and isothermal titration calorimetry, we first confirmed the binding between NpmA and 30S. Next, we performed amide Hydrogen/Deuterium Exchange Mass Spectrometry (HDXMS) of apo NpmA and in the presence and absence of SAM/SAH. We observed that ligand binding resulted in time-dependent differences in deuterium exchange not only at the ligand-binding pocket (D25-D55 and A86-E112) but also in distal regions (F62-F82 and Y113-S144) of NpmA. These results provide insights into methylation group donor cofactor-mediated allostery in NpmA in the ligand-bound states, which could not be observed in the static endpoint crystal structures. We predict that the two distal sites in NpmA form part of the allosteric sites that importantly are part of the main 16S rRNA binding interface. Thus HDXMS helped uncover allosteric communication relays that couple SAM/SAH binding sites with the ribosome-binding site. This highlights how HDXMS together with X-ray crystallography can provide important allosteric insights in protein-ligand complexes. © 2016 The Author(s).
Source Title: Scientific Reports
URI: https://scholarbank.nus.edu.sg/handle/10635/173978
ISSN: 20452322
DOI: 10.1038/srep37061
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