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Insights in Enzyme Research
ISSN: 2573-4466
E u r o S c i C o n C o n g r e s s o n
Enzymology and
Molecular Biology
A u g u s t 1 3 - 1 4 , 2 0 1 8
P a r i s , F r a n c e
Enzymology 2018
P
rotein dynamics manifested through structural flexibility play a central role in the function of biological molecules. Here
we explore the substrate-mediated change in protein flexibility of an enzyme antibiotic target,
Clostridium botulinum
dihydrodipicolinate synthase (DHDPS). We demonstrate that the substrate, pyruvate, stabilizes the more active dimer-of-dimers
or tetrameric form of the enzyme. Surprisingly, there is little difference between the crystal structures of apo and substrate-
bound DHDPS, suggesting protein dynamics may be important. Neutron and small angle X-ray scattering experiments were
used to probe substrate-induced dynamics on the sub-second timescale, but no significant changes were observed. We have
therefore developed a simple technique, coined Protein Dynamics-Mass Spectrometry (ProD-MS), which enables measurement
of time-dependent alkylation of cysteine residues. ProD-MS together with X-ray crystallography and analytical ultracentrifugation
analyses indicates that pyruvate locks the conformation of the dimer that promotes docking to the more active tetrameric form,
offering new insight into ligand-mediated stabilization of multimeric enzymes
.
M.Perugini@latrobe.edu.auSubstrate locking promotes dimer-dimer docking
of an enzyme antibiotic target
Matthew A Perugini
1,2
, Sarah C Atkinson
1,2,3
, Con Dogovski
2
,
Kathleen Wood
4
, Michael D W Griffin
2
, Michael A Gorman
5
, Lil-
ian Hor
1
, Cyril F Reboul
3
, Ashley M Buckle
3
, Joachim Wuttke
6
,
Michael W Parker
2,5
and Renwick C J Dobson
7
1
La Trobe University, Melbourne, Australia
2
Bio21 Institute, University of Melbourne, Victoria, Australia
3
Monash University, Clayton, Australia
4
Australian Nuclear Science & Technology Organisation, New South Wales, Australia
5
St. Vincent's Institute of Medical Research, Victoria, Australia
6
JCNS-Heinz Maier-Leibnitz Zentrum (MLZ), Forschungszentrum Juelich GmbH, Garching, Germany
7
University of Canterbury, Christchurch, New Zealand
Insights Enzyme Res 2018, Volume 2
DOI: 10.21767/2573-4466-C1-003