10.5061/DRYAD.031SF
Dalla, Safaa
University of Hamburg
Baum, Michael
University of Hamburg
Dobler, Susanne
University of Hamburg
Data from: Substitutions in the cardenolide binding site and interaction
of subunits affect kinetics besides cardenolide sensitivity of insect
Na,K-ATPase
Dryad
dataset
2017
Oncopeltus fasciatus
K-ATPase
Amino acid substitutions
Subunit interaction
Cardenolide resistance
2017-09-06T20:02:46Z
2017-09-06T20:02:46Z
en
https://doi.org/10.1016/j.ibmb.2017.08.005
78071 bytes
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CC0 1.0 Universal (CC0 1.0) Public Domain Dedication
Substitutions within the cardenolide target site of several insects'
Na,K-ATPase α-subunits may confer resistance against toxic cardenolides.
However, to which extent these substitutions alter the Na,K-ATPase's
kinetic properties and how they interact with different β-subunits is not
clear. The cardenolide-adapted milkweed bug Oncopeltus fasciatus possesses
three paralogs of the α-subunit (A, B, and C) that differ in number and
identity of resistance-conferring substitutions. We introduced these
substitutions into the α-subunit of Drosophila melanogaster and combined
them with the β-subunits Nrv2.2 and Nrv3. The substitutions
Q111T-N122H-F786N-T797A (A-copy mimic) and Q111T-N122H-F786N (B-copy
mimic) mediated high insensitivity to ouabain, yet they drastically
lowered ATPase activity. Remarkably, the identity of the β-subunit was
decisive and all α-subunits were less active when combined with Nrv3 than
when combined with Nrv2.2. Both the substitutions and the co-expressed
β-subunit strongly affected the enyzme's affinity for Na+ and K+. Na+
affinity was considerably higher for all enzymes expressed with nrv3 while
expression with nrv2.2 increased K+ affinity. Our results provide the
first evidence that resistance against cardenolides comes at the cost of
significantly altered kinetic properties of the Na,K-ATPase. The β-subunit
can strongly modulate these properties but cannot fully compensate for the
effect of the substitutions.
Raw data Na affinity Nrv2.2-alphasData on Na+ affinity of Na,K-ATPases
consisting of mutant and wildtype D. melanogaster alpha subunits and the
beta subunit Nrv2.2Raw data Na affinity Nrv3-alphasData on Na+ affinity of
Na,K-ATPases consisting of mutant and wildtype D. melanogaster alpha
subunits and the beta subunit Nrv3Raw data K affinity Nrv2.2-alphasData on
K+ affinity of Na,K-ATPases consisting of mutant and wildtype D.
melanogaster alpha subunits and the beta subunit Nrv2.2Raw data K affinity
Nrv3-alphasData on K+ affinity of Na,K-ATPases consisting of mutant and
wildtype D. melanogaster alpha subunits and the beta subunit Nrv3Raw Data
ATP-affinity Nrv2.2-alphasData on ATP affinity of Na,K-ATPases consisting
of mutant and wildtype D. melanogaster alpha subunits and the beta subunit
Nrv2.2Raw Data ATP-affinity Nrv3-alphasData on ATP affinity of
Na,K-ATPases consisting of mutant and wildtype D. melanogaster alpha
subunits and the beta subunit Nrv3