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@ARTICLE{Kaschner:829442,
author = {Kaschner, Marco and Schillinger, Oliver and Fettweiss, Timo
and Nutschel, Christina and Krause, Frank and Fulton,
Alexander and Strodel, Birgit and Stadler, Andreas and
Jaeger, Karl-Erich and Krauss, Ulrich},
title = {{A} combination of mutational and computational scanning
guides the design of an artificial ligand-binding controlled
lipase},
journal = {Scientific reports},
volume = {7},
issn = {2045-2322},
address = {London},
publisher = {Nature Publishing Group},
reportid = {FZJ-2017-03146},
pages = {42592 -},
year = {2017},
abstract = {Allostery, i.e. the control of enzyme activity by a small
molecule at a location distant from the enzyme’s active
site, represents a mechanism essential for sustaining life.
The rational design of allostery is a non-trivial task but
can be achieved by fusion of a sensory domain, which
responds to environmental stimuli with a change in its
structure. Hereby, the site of domain fusion is difficult to
predict. We here explore the possibility to rationally
engineer allostery into the naturally not allosterically
regulated Bacillus subtilis lipase A, by fusion of the
citrate-binding sensor-domain of the CitA sensory-kinase of
Klebsiella pneumoniae. The site of domain fusion was
rationally determined based on whole-protein site-saturation
mutagenesis data, complemented by computational
evolutionary-coupling analyses. Functional assays, combined
with biochemical and biophysical studies suggest a mechanism
for control, similar but distinct to the one of the parent
CitA protein, with citrate acting as an indirect modulator
of Triton-X100 inhibition of the fusion protein. Our study
demonstrates that the introduction of ligand-dependent
regulatory control by domain fusion is surprisingly facile,
suggesting that the catalytic mechanism of some enzymes may
be evolutionary optimized in a way that it can easily be
perturbed by small conformational changes.},
cin = {ICS-1 / Neutronenstreuung ; JCNS-1 / ICS-6 / IMET},
ddc = {000},
cid = {I:(DE-Juel1)ICS-1-20110106 / I:(DE-Juel1)JCNS-1-20110106 /
I:(DE-Juel1)ICS-6-20110106 / I:(DE-Juel1)IMET-20090612},
pnm = {551 - Functional Macromolecules and Complexes (POF3-551) /
6G4 - Jülich Centre for Neutron Research (JCNS) (POF3-623)
/ 6215 - Soft Matter, Health and Life Sciences (POF3-621) /
581 - Biotechnology (POF3-581)},
pid = {G:(DE-HGF)POF3-551 / G:(DE-HGF)POF3-6G4 /
G:(DE-HGF)POF3-6215 / G:(DE-HGF)POF3-581},
typ = {PUB:(DE-HGF)16},
UT = {WOS:000394417100001},
doi = {10.1038/srep42592},
url = {https://juser.fz-juelich.de/record/829442},
}