Аннотация
Activation of the cardiac ryanodine receptor (RyR2) by Ca(2)+ is an
essential step in excitation-contraction coupling in heart muscle.
However, little is known about the molecular basis of activation
of RyR2 by Ca(2)+. In this study, we investigated the role in Ca(2)+
sensing of the conserved glutamate 3987 located in the predicted
transmembrane segment M2 of the mouse RyR2. Single point mutation
of this conserved glutamate to alanine (E3987A) reduced markedly
the sensitivity of the channel to activation by Ca(2)+, as measured
by using single-channel recordings in planar lipid bilayers and by
(3)Hryanodine binding assay. However, this mutation did not alter
the affinity of (3)Hryanodine binding and the single-channel conductance.
In addition, the E3987A mutant channel was activated by caffeine
and ATP, was inhibited by Mg(2)+, and was modified by ryanodine in
a fashion similar to that of the wild-type channel. Coexpression
of the wild-type and mutant E3987A RyR2 proteins in HEK293 cells
produced individual single channels with intermediate sensitivities
to activating Ca(2)+. These results are consistent with the view
that glutamate 3987 is a major determinant of Ca(2)+ sensitivity
to activation of the mouse RyR2 channel, and that Ca(2)+ sensing
by RyR2 involves the cooperative action between ryanodine receptor
monomers. The results of this study also provide initial insights
into the structural and functional properties of the mouse RyR2,
which should be useful for studying RyR2 function and regulation
in genetically modified mouse models.
- 11429443
- acid,
- adenosine
- alanine,
- animals,
- caffeine,
- calcium
- calcium,
- channel
- channel,
- co,
- dose-response
- drug,
- electrophysiology,
- gating,
- gl,
- glutamic
- gov't,
- heart,
- humans,
- ion
- magnesium,
- mice,
- muscle
- mutation,
- non-u.s.
- ntraction,
- point
- receptor
- relationship,
- release
- research
- reticulum,
- ryanodine
- ryanodine,
- sarcoplasmic
- support,
- transfection,
- triphosphate,
- utamic
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