Gβγ binding increases the open time of I(KACh): Kinetic evidence for multiple Gβγ binding sites

I(KACh) is an inwardly rectifying potassium channel that plays an important role in the regulation of mammalian heart rate. I(KACh) is activated by direct interaction with Gβγ subunits of pertussis toxin- sensitive heterotrimeric G-proteins. The stoichiometry of the Gβγ/channel complex is currently unknown, and kinetic analysis of the channel behavior has led to conflicting conclusions. Here, we analyze the kinetics of the native I(KACh) channel in inside-out cardiomyocyte patches activated directly by Gβγ. We conclude that the channel has at least two open states and that binding of Gβγ, prolongs its mean open time duration. These findings imply the existence of at least two binding sites on the channel complex for Gβγ. We also show that the duration of the channel opening is negatively correlated with the duration of subsequent channel closing, which further constrains the possible kinetic models. A simple qualitative model describing the kinetic behavior of I(KACh) is presented.