SCH28080, a K+-competitive inhibitor of the gastric H,K-ATPase, binds near the M5-6 luminal loop, preventing K+ access to the ion binding domain.
SCH28080, a K+-competitive inhibitor of the gastric H,K-ATPase, binds near the M5-6 luminal loop, preventing K+ access to the ion binding domain.
复制标题
SCH28080 是胃 H,K-ATP 酶的 K 竞争性抑制剂，结合在 M5-6 腔环附近，阻止 K 进入离子结合域。
作者：
Vagin,O;Denevich,S;Munson,K;Sachs,G
咪唑并[1，2-α]吡啶SCH28080对胃H，K-ATPase的抑制作用与K+或其替代物NH4+是严格竞争的。Y799F、Y802F、I803L、S806N、V807I(M5)、L811V(M5−6)、Y928H(M8)和Q905N(M7−8)突变对抑制动力学没有影响，突变P798C、Y802L、P810A、P810G、C813A或S、I814V或-F、F818C、T823V(M5、M5−6和M6)、E914Q、F917Y、G918E、T929L和Q905N(M7−8)对抑制动力学没有影响。而F932L(M7、−8和M8)在不影响动力学性质的情况下，对SCH28080的亲和力降低了10倍。相反，M5和M6之间环中的L809F替换导致∼抑制剂亲和力降低100倍，L809V、I816L、Y925F和M937V(M5−6、M6和M8)替换使抑制剂亲和力降低10倍，所有这些都导致非竞争性动力学。突变体L811F、Y922I和I940A也使抑制剂亲和力降低了10倍，但导致了混合抑制。突变I819L、Q923V和Y925A也表现出混合抑制，但抑制物亲和力没有变化。这些数据以及C813T突变体中SCH28080亲和力的9倍损失表明，SCH28080的结合域包含在M5−6环的L809和M6管腔末端的C813之间的表面，大约两个螺旋向下的离子结合区，在那里它阻止了正常的离子访问途径。根据E2构象中Ca-−酶的模型(pdb条目1kju)，改变动力学性质的突变体排列在M8的一侧和M5 ATPase 6环和M6本身的相邻侧。这表明该区域的突变改变了酶的结构，使得K+即使与SCH28080结合也可以访问离子结合区域。
Inhibition of the gastric H,K-ATPase by the imidazo[1,2-α]pyridine, SCH28080, is strictly competitive with respect to K+or its surrogate, NH4+. The inhibitory kinetics [Vmax,Km,app(NH4+),Ki(SCH28080), and competitive, mixed, or noncompetitive] of mutants can define the inhibitor binding domain and the route to the ion binding region within M4−6. While mutations Y799F, Y802F, I803L, S806N, V807I (M5), L811V (M5−6), Y928H (M8), and Q905N (M7−8) had no effect on inhibitor kinetics, mutations P798C, Y802L, P810A, P810G, C813A or -S, I814V or -F, F818C, T823V (M5, M5−6, and M6), E914Q, F917Y, G918E, T929L, and F932L (M7−8 and M8) reduced the affinity for SCH28080 up to 10-fold without affecting the nature of the kinetics. In contrast, the L809F substitution in the loop between M5 and M6 resulted in an ∼100-fold decrease in inhibitor affinity, and substitutions L809V, I816L, Y925F, and M937V (M5−6, M6, and M8) reduced the inhibitor affinity by 10-fold, all resulting in noncompetitive kinetics. The mutants L811F, Y922I, and I940A also reduced the inhibitor affinity up to 10-fold but resulted in mixed inhibition. The mutations I819L, Q923V, and Y925A also gave mixed inhibition but without a change in inhibitor affinity. These data, and the 9-fold loss of SCH28080 affinity in the C813T mutant, suggest that the binding domain for SCH28080 contains the surface between L809 in the M5−6 loop and C813 at the luminal end of M6, approximately two helical turns down from the ion binding region, where it blocks the normal ion access pathway. On the basis of a model of the Ca-ATPase in the E2conformation (PDB entry 1kju), the mutants that change the nature of the kinetics are arranged on one side of M8 and on the adjacent side of the M5−6 loop and M6 itself. This suggests that mutations in this region modify the enzyme structure so that K+can access the ion binding domain even with SCH28080 bound.