Histidine(118) in the S2-S3 linker specifically controls activation of the KAT1 channel expressed in Xenopus oocytes.
The guard cell K(+) channel KAT1, cloned from Arabidopsis thaliana, is activated by hyperpolarization and regulated by a variety of physiological factors. Low internal pH accelerated the activation kinetics of the KAT1 channel expressed in Xenopus oocytes with a pK of approximately 6, similar to guard cells in vivo. Mutations of histidine-118 located in the putative cytoplasmic linker between the S2 and S3 segments profoundly affected the gating behavior and pH dependence. At pH 7.2, substitution with a negatively charged amino acid (glutamate, aspartate) specifically slowed the activation time course, whereas that with a positively charged amino acid (lysine, arginine) accelerated. These mutations did not alter the channel's deactivation time course or the gating behavior after the first opening. Introducing an uncharged amino acid (alanine, asparagine) at position 118 did not have any obvious effect on the activation kinetics at pH 7.2. The charged substitutions markedly decreased the sensitivity of the KAT1 channel to internal pH in the physiological range. We propose a linear kinetic scheme to account for the KAT1 activation time course at the voltages where the opening transitions dominate. Changes in one forward rate constant in the model adequately account for the effects of the mutations at position 118 in the S2-S3 linker segment. These results provide a molecular and biophysical basis for the diversity in the activation kinetics of inward rectifiers among different plant species.
Tang
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XD
Department of Physiology and Biophysics, The University of Iowa, Iowa City, Iowa 52242, USA.
Marten
I
I
Dietrich
P
P
Ivashikina
N
N
Hedrich
R
R
Hoshi
T
T
D000818:Animals; D017360:Arabidopsis / Q000235:genetics / Q000502:physiology; D029681:Arabidopsis Proteins; D001665:Binding Sites; D003001:Cloning, Molecular; D005260:Female; D006639:Histidine; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D015640:Ion Channel Gating; D008564:Membrane Potentials / Q000502:physiology; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D010940:Plant Proteins / Q000502:physiology; D015221:Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D024661:Potassium Channels, Inwardly Rectifying; D017433:Protein Structure, Secondary; D011930:Reaction Time; D014982:Xenopus laevis
D029681:Arabidopsis Proteins; C501210:KAT1 protein, Arabidopsis; D010940:Plant Proteins; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D006639:Histidine