Activation and inhibition of K-ATP currents by guanine nucleotides is mediated by different channel subunits.
The ATP-sensitive potassium channel (K-ATP channel) plays a key role in insulin secretion from pancreatic beta-cells. It is closed by glucose metabolism, which stimulates secretion, and opened by the drug diazoxide, which inhibits insulin release. Metabolic regulation is mediated by changes in ATP and MgADP concentration, which inhibit and potentiate channel activity, respectively. The beta-cell K-ATP channel consists of a pore-forming subunit, Kir6.2, and a regulatory subunit, SUR1. The site at which ATP mediates channel inhibition lies on Kir6.2, while the potentiatory action of MgADP involves the nucleotide-binding domains of SUR1. K-ATP channels are also activated by MgGTP and MgGDP. Furthermore, both nucleotides support the stimulatory actions of diazoxide. It is not known, however, whether guanine nucleotides mediate their effects by direct interaction with one or more of the K-ATP channel subunits or indirectly via a GTP-binding protein. We used a truncated form of Kir6.2, which expresses independently of SUR1, to show that GTP blocks K-ATP currents by interaction with Kir6.2 and that the potentiatory effects of GTP are endowed by SUR1. We also showed that mutation of the lysine residue in the Walker A motif of either the first (K719A) or second (K1384M) nucleotide-binding domain of SUR1 abolished both the potentiatory effects of GTP and GDP on K-ATP currents and their ability to support stimulation by diazoxide. This argues that the stimulatory effects of guanine nucleotides require the presence of both Walker A lysines.
Trapp
S
S
University Laboratory of Physiology, Parks Road, Oxford OX1 3PT, United Kingdom.
Tucker
S J
SJ
Ashcroft
F M
FM
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D015640:Ion Channel Gating / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D014981:Xenopus
D015221:Potassium Channels; D000255:Adenosine Triphosphate