Direct Regulation of Prokaryotic Kir Channel by Cholesterol

Abstract

Our earlier studies have shown that channel activity of Kir2 subfamily of inward rectifiers is strongly suppressed by the elevation of cellular cholesterol. The goal of this study is to determine whether cholesterol suppresses Kir channels directly. To achieve this goal, purified prokaryotic Kir (KirBac1.1) channels were incorporated into liposomes of defined lipid composition, and channel activity was assayed by 86 ple sterols suggests that cholesterol-induced inhibition of KirBac1.1 channels is mediated by specific interactions rather than by changes in the physical properties of the lipid bilayer. In contrast to the inhibition of KirBac1.1 activity, cholesterol had no effect on the activity of reconstituted KscA channels (at up to 250 g/mg of phospholipid). Taken together, these observations demonstrate that cholesterol suppresses Kir channels in a pure protein-lipid environment and suggest that the interaction is direct and specific.

Enantioselective Protein-Sterol Interactions Mediate Regulation of Both Prokaryotic and Eukaryotic Inward Rectifier K+ Channels by Cholesterol

“…By incorporating these channels into liposomes, previous studies have shown that these proteins form functional K + channels [24]–[26], [30], [32], [35], and that KirBac1.1-mediated 86 Rb + flux is readily suppressed by the addition of cholesterol to 9∶1 POPE∶POPG liposomes [21]. Here we confirm these results (Fig.…”

Identification of a Binding Motif in the S5 Helix That Confers Cholesterol Sensitivity to the TRPV1 Ion Channel

“…In addition, in recent years compelling evidence has shown a specific interaction between cholesterol and several protein channels (13,15). That is, based on experiments with structural analogs of cholesterol and channels with specific point mutations, it has been found that cholesterol binds to certain channel proteins, including some with structural similarities to TRPV1 (16). For either of these indirect or direct mechanisms, cholesterol modifies the energy difference between the open and closed states of the channel.…”

“…First, epicholesterol (␣-3-OH-cholesterol epimeric form) was substituted for cholesterol (see Fig. 1), as previously done with other channels (16,19). Second, capsaicin-induced currents were measured for membranes transfected with rTRPV1s with specific point mutations in the S5 transmembrane helix, which has a sequence consistent with the cholesterol recognition amino acid consensus (CRAC) sequence found in several transmembrane proteins that bind cholesterol (20,21).…”

Acute Cholesterol-induced Anti-natriuretic Effects

“…It is established that membrane cholesterol content can modulate the activity of integral membrane proteins and that experimental depletion or enrichment of cholesterol modifies channel and transporter activity (1,(3)(4)(5)(6)(7)(8). These effects have been attributed to either direct interaction between cholesterol and the membrane channels and transporters (8) or changes of membrane order and fluidity brought about by changes of lipid composition (9).…”

“…These effects have been attributed to either direct interaction between cholesterol and the membrane channels and transporters (8) or changes of membrane order and fluidity brought about by changes of lipid composition (9). A contributor to both mechanisms is the presence of cholesterol-rich "raft" membrane microdomains, where integral proteins can preferentially partition and where partitioning into such domains can modify protein activity and/or stability (10 -14).…”