Differential pyridine nucleotide coenzyme binding to the beta-subunit of the voltage-sensitive K+ channel: a mechanism for redox regulation of excitability?

The pore-forming subunits of the voltage-sensitive K(+) channel (K(v)) associate with ancillary beta-subunits that regulate inactivation and voltage-dependence of the channel. The beta-subunits are members of the aldo-keto reductase (AKR) superfamily. We have previously demonstrated that recombinant K(v)beta2.1 displays tight binding to NADP(H). The protein also binds NAD(H), but with less affinity. To assess the physiological significance of this binding, we examined how pyridine nucleotides regulate the K(v)beta-mediated inactivation of K(+) channels. Transient transfection of COS-7 cells with an pIRES-hrGFP vector containing the Kvalpha1.5 cDNA led to the appearance of the K(v)1.5 protein in the membrane fraction and large non-inactivating potassium currents were recorded from the transfected cells. No such currents were observed in cells transfected with the empty vector alone or with K(v)beta1.3 (AKR6A3), which was localized to the cytoplasm. In contrast, K(v)beta1.3 co-transfected with Kvalpha1.5 was localized to the membrane, suggesting high affinity binding of the two proteins. Moreover, the K currents recorded from cells transfected with both K(v)alpha1.5 and K(v)beta1.3 displayed pronounced inactivation. Inclusion of 1 mM NAD(+) in the internal solution of the patch pipette abolished K(v)beta-induced inactivation of K(v)1.5 currents, but did not affect the non-inactivating currents recorded from cells transfected with K(v)1.5 alone, indicating that in the absence of K(v)beta, NAD(+) does not affect the activity of K(v)alpha. The inactivating currents recorded from cells expressing both K(v)alpha1.5 and K(v)beta1.3 were unaffected by the inclusion of 0.1 mM NADPH in the pipette solution. Together, these data suggest that NADPH and NAD(+) impart different conformational states to the K(v)beta protein and that only the NADPH bound K(v)beta imparts inactivation to non-inactivating K(+) currents. Thus, differential binding of pyridine nucleotide coenzymes to K(v)beta could regulate membrane potential and excitability as a function of the cellular redox state. Because NAD(+)/NADPH ratio is sensitive to oxygen concentration, the differential changes in K(v)beta-mediated inactivation of K currents by NAD(+) and NADPH could represent an oxygen-sensing mechanism.