Cox D H, Aldrich R W
Molecular Cardiology Research Institute, New England Medical Center, Boston 02111, USA.
J Gen Physiol. 2000 Sep;116(3):411-32. doi: 10.1085/jgp.116.3.411.
Over the past few years, it has become clear that an important mechanism by which large-conductance Ca(2+)-activated K(+) channel (BK(Ca)) activity is regulated is the tissue-specific expression of auxiliary beta subunits. The first of these to be identified, beta1, is expressed predominately in smooth muscle and causes dramatic effects, increasing the apparent affinity of the channel for Ca(2+) 10-fold at 0 mV, and shifting the range of voltages over which the channel activates -80 mV at 9.1 microM Ca(2+). With this study, we address the question: which aspects of BK(Ca) gating are altered by beta1 to bring about these effects: Ca(2+) binding, voltage sensing, or the intrinsic energetics of channel opening? The approach we have taken is to express the beta1 subunit together with the BK(Ca) alpha subunit in Xenopus oocytes, and then to compare beta1's steady state effects over a wide range of Ca(2+) concentrations and membrane voltages to those predicted by allosteric models whose parameters have been altered to mimic changes in the aspects of gating listed above. The results of our analysis suggest that much of beta1's steady state effects can be accounted for by a reduction in the intrinsic energy the channel must overcome to open and a decrease in its voltage sensitivity, with little change in the affinity of the channel for Ca(2+) when it is either open or closed. Interestingly, however, the small changes in Ca(2+) binding affinity suggested by our analysis (K(c) 7.4 microM --> 9.6 microM; K(o) = 0.80 microM --> 0.65 microM) do appear to be functionally important. We also show that beta1 affects the mSlo conductance-voltage relation in the essential absence of Ca(2+), shifting it +20 mV and reducing its apparent gating charge 38%, and we develop methods for distinguishing between alterations in Ca(2+) binding and other aspects of BK(Ca) channel gating that may be of general use.
在过去几年中，已明确大电导钙激活钾通道（BK(Ca)）活性受调节的一个重要机制是辅助β亚基的组织特异性表达。其中第一个被鉴定的β1，主要在平滑肌中表达，并产生显著影响，在0 mV时将通道对Ca(2+)的表观亲和力提高10倍，并在9.1 microM Ca(2+)时将通道激活的电压范围向负向移动80 mV。通过本研究，我们探讨了以下问题：β1改变了BK(Ca)门控的哪些方面从而产生这些效应：Ca(2+)结合、电压传感还是通道开放的内在能量学？我们采用的方法是在非洲爪蟾卵母细胞中共同表达β1亚基和BK(Ca)α亚基，然后在广泛的Ca(2+)浓度和膜电压范围内，将β1的稳态效应与通过改变参数以模拟上述门控方面变化的变构模型所预测的效应进行比较。我们的分析结果表明，β1的许多稳态效应可以通过通道开放时必须克服的内在能量降低以及电压敏感性降低来解释，而通道在开放或关闭时对Ca(2+)的亲和力变化很小。然而，有趣的是，我们的分析表明Ca(2+)结合亲和力的微小变化（K(c) 7.4 microM --> 9.6 microM；K(o) = 0.80 microM --> 0.65 microM）似乎在功能上很重要。我们还表明，在基本没有Ca(2+)的情况下，β1会影响mSlo电导 - 电压关系，使其正向移动20 mV并将其表观门控电荷降低38%，并且我们开发了区分Ca(2+)结合变化和BK(Ca)通道门控其他方面变化的方法，这些方法可能具有普遍用途。
