质膜Ca2+-ATPase的结构与功能

质膜Ca2+-ATPase (PMCA)是P型ATPase家族的一员,在真核细胞中主要负责信号刺激后胞内高浓度Ca2+的清除扫尾工作,并对维持静息状态下较低Ca2+浓度起着重要的调节作用.PMCA的一级结构已被确定,拓扑学结构显示,它有10个跨膜区和3个胞浆功能区.它的4个编码基因可产生4种亚型(PMCA 1~4),这些亚型在功能与分布上存在差异.PMCA的活性可被钙调蛋白等多种因素调节,这与其结构特征息息相关.近年来,PMCA已被证实与脂筏结构有一定关联,它在信号传导和细胞凋亡中的作用也成为目前科学研究的焦点.本文主要对PMCA的结构、亚型和功能的研究现状进行综述.     

高温胁迫对花生幼苗光合速率、叶绿素含量、叶绿体Ca2+-ATPase、Mg2+-ATPase及Ca2+分布的影响

将水培后盆栽的花生幼苗,置于培养箱42℃高温培养,定时测定幼苗叶光合速率、叶绿素含量和叶绿体Ca2 -ATPase、Mg2 -ATPase的相对活性,并观察幼叶细胞内Ca2 分布的变化。试验结果表明:高温胁迫过程中,光合速率及叶绿素含量都随处理时间的延伸而下降,并呈显著正相关;叶绿体Ca2 -ATPase和Mg2 -ATPase高温胁迫过程中相对活性呈先升后降趋势,Ca2 -ATPase热敏性高于Mg2 -ATPase;高温胁迫过程中,Ca2 具有从胞外转运到胞质内和叶绿体中的趋势,Ca2 能够稳定高温胁迫下叶肉细胞膜和叶绿体的超微结构。     

高温胁迫对花生幼苗光合速率、叶绿素含量、叶绿体Ca2+-ATPase、Mg2+-ATPase及Ca2+分布的影响

将水培后盆栽的花生幼苗,置于培养箱42℃高温培养,定时测定幼苗叶光合速率、叶绿素含量和叶绿体Ca^2＋-ATPase、Mg^2＋-ATPase的相对活性,并观察幼叶细胞内Ca^2＋分布的变化。试验结果表明：高温胁迫过程中,光合速率及叶绿素含量都随处理时间的延伸而下降,并呈显著正相关;叶绿体Ca^2＋-ATPase和Mg^2＋-ATPase高温胁迫过程中相对活性呈先升后降趋势,Ca^2＋-ATPase热敏性高于Mg^2＋-ATPase;高温胁迫过程中,Ca^2＋具有从胞外转运到胞质内和叶绿体中的趋势,Ca^2＋能够稳定高温胁迫下叶肉细胞膜和叶绿体的超微结构。     

稀土离子对CaM及Ca2+-Mg2+-ATPase活力及CD研究

研究了稀土离子（Ｌｎ３＋）对钙调蛋白（ＣａＭ）调控的Ｃａ２＋－Ｍｇ２＋－ＡＴＰａｓｅ的活力影响。结果表明,在ＣａＭ和Ｃａ２＋－Ｍｇ２＋－ＡＴＰａｓｅ的体系中,一些Ｌｎ３＋（Ｌａ３＋、Ｇｄ３＋）对由ＣａＭ调节的Ｃａ２＋－Ｍｇ２＋－ＡＴＰａｓｅ的活力影响呈现双相效应,即Ｌｎ３＋在低浓度时,能提高激活Ｃａ２＋－Ｍｇ２＋－ＡＴＰａｓｅ的水解活力;在高浓度时,则抑制ＣａＭ调节Ｃａ２＋－Ｍｇ２＋－ＡＴＰａｓｅ活力的能力;少数Ｌｎ３＋（Ｓｍ３＋）仅表现出抑制效应。在无ＣａＭ的Ｃａ２＋－Ｍｇ２＋－ＡＴＰａｓｅ体系中,高浓度的Ｌｎ３＋抑制Ｃａ２＋－Ｍｇ２＋－ＡＴＰａｓｅ的基础活力。结合圆二色（ＣＤ）谱信息对Ｌｎ３＋和ＣａＭ相互作用的分子机制进行了初步的探讨。     

番茄碱研究进展以及对鸡红细胞膜Ca2+-Mg2+-ATPase活性的影响

综述了番茄碱的研究进展,并且研究了番茄碱对鸡红细胞膜Ca2 -Mg2 -ATPase的影响,实验结果表明:当番茄碱的浓度在0-1 mmol／L时,随着番茄碱浓度的增加,Ca2 -Mg2 -ATPase的活性呈下降趋势。为进一步研究开发番茄碱奠定了基础。     

胃粘膜Na~+-K~+-Mg~(2+)-ATPase在适应性细胞保护机制中的作用

本文观察了胃粘膜(Na~+-K~+-Mg~(2+))-ATPase在适应性细胞保护机制中的作用,并分析了其与内源性PG的关系。结果表明,哇巴因(一种(Na~+-K~+-Mg~(2+))-ATPase的抑制剂)可部分抑制胃蛋白酶150U(溶于0.1mol/L盐酸中)和20％乙醇的适应性细胞保护作用,并呈现明显的量效关系。用上述两种弱刺激灌胃后15min,胃粘膜(Na~+-K~+-Mg~(2+))-ATPase活力明显升高,也呈现明显的量效关系。预先给予消炎痛以抑制内源性PG的合成,则可阻断弱刺激所诱发的胃粘膜(Na~+-K~+-Mg~(2+))-ATPase活力的升高;若在此基础上再给予外源性PGE_2,又可解除消炎痛的阻断作用。这些结果说明,弱刺激通过内源性PG,进而促进胃粘膜(Na~+-K~+-Mg~(2+))-ATPase活力升高,使粘膜抵抗损伤的能力增强,可能是其保护作用的重要机制之一。     

心肌缺血预处理后Ca2+*Mg2+-ATPase及SDH活性的变化

198 6年Ｍｕｒｒｙ等首次发现实验狗心肌在经历了短暂性缺血后产生对随后持续严重缺血再灌注的保护作用 ,并称此现象为“缺血预处理 (ｉｓｃｈｅｍｉｃｐｒｏｅｏｎｄｉｔｉｏｎｉｎｇ,ＩＰＣ)。ＩＰＣ现象的发现为心肌保护提供了一条新的途径。实验及临床研究均发现其可以限制心肌梗塞范围 ,增加心肌收缩力 ,降低再灌性心律失常 ,但心肌的这一内源性保护机制不清。有人认为ＩＰＣ的产生机制与纠正细胞内外的离子紊乱和心肌能量代谢有关。Ｃａ2 ·Ｍｇ2 ＡＴＰａｓｅ在维持细胞内外的正常离子浓度中起重要作用 ,琥珀酸脱氢酶 (ＳＤＨ)是…     

草莓果实成熟过程中细胞Ca2+-ATPase活性的变化

‘春星’草莓果实成熟时,总糖和花青苷含量增加,呼吸速率也显著升高;同时,细胞溶质Ca2 -ATPase活性和微粒体膜的Ca2 ．ATPase总活性变化具有相似的特点,即先升高,至粉红期达到高峰,全红期又下降,在微粒体膜中以质膜Ca2 -ATPase占的比例最高。抑制质膜Ca2 -ATPase活性的Na3VO4能促进草莓果实花青苷积累、降低可溶性总糖含量,但对呼吸速率的影响则因草莓果实成熟度不同而异。     

空泡膜类型H~+-ATPase的研究进展

真核细胞内空泡细胞器,如高尔基体、内质网、溶酶体等,膜上存在的质子泵ATPase 与线粒体类型的质子泵 ATPase 类似.近几年对该类型 H~+-ATPase 的结构、作用机制进行了深入的研究,证明这是一类新型质子泵,在进化的过程中与线粒体类型的 H~+-ATPase 有密切的亲缘关系.     

草莓果实成熟过程中细胞Ca2+-ATPase活性的变化

‘春星’草莓果实成熟时,总糖和花青苷含量增加,呼吸速率也显著升高;同时,细胞溶质Ca2+-ATPase活性和微粒体膜的Ca2+-ATPase总活性变化具有相似的特点,即先升高,至粉红期达到高峰,全红期又下降,在微粒体膜中以质膜Ca2+-ATPase占的比例最高。抑制质膜Ca2+-ATPase活性的Na3VO4能促进草莓果实花青苷积累、降低可溶性总糖含量,但对呼吸速率的影响则因草莓果实成熟度不同而异。     

植物质膜H+-ATPase的研究进展

质膜H -ATPase参与植物细胞的物质跨膜转运、细胞的伸长生长、气孔的开闭以及植物对环境胁迫的响应等生理过程,是植物生命活动的“主宰酶”。其活性调节涉及激素、环境因子等多种因素,可发生在转录、翻译和酶分子等多级水平。因此,在植物生长发育过程中,质膜H -ATPase活性的调节对生理活动起重要作用。本文就植物质膜H -ATPase的结构特征、生理功能、活性变化及其调节机理等的研究进展进行综述,以进一步揭示该酶的生理功能及其调节机理与植物生命活动过程的关系。     

α-adrenergic receptor regulation of Ca2+/Mg2+-ATPase in brain synaptic membranes

The effects of ₁ and ₂ receptor ligands on Ca²⁺/Mg²⁺-ATPase have been studied using synaptosomal plasma membranes isolated from rat brain cortex. Both phenylephrine and clonidine inhibited Ca²⁺/Mg²⁺-ATPase, in a concentration-dependent fashion. IC₅₀ values for half-maximal inhibition for phenylephrine and clonidine were 29 M and 18 M, respectively. The inhibitory effect of phenylephrine was reversed by the alpha antagonist prazosin while yohimbine and rauwolscine reversed the inhibition of enzyme activity by clonidine. The two antagonist subtypes were effective only against the respective agonist subtypes, demonstrating distinct subtype preferences. Analysis of the kinetics of enzyme inhibition indicate both agonists to be noncompetitive. Some evidence suggests that yohimbine may exhibit mixed agonist/antagonist properties which depend on [Ca²⁺]. The present study provides biochemical evidence to support auto receptor adrenergic receptor regulation of neurotransmitter release.     

NaHCO3胁迫下星星草根中Ca2+与Ca2+-ATPase 的超微细胞化学定位

利用焦锑酸盐和磷酸铅沉淀技术分别对NaHCO₃胁迫条件下星星草(Puccinellia tenuiflora)根中Ca²⁺和Ca²⁺-ATPase 进行超微细胞化学定位研究, 旨在进一步探讨Ca²⁺在NaHCO₃胁迫诱导胞内信号转导过程中的作用, 以及Ca²⁺-ATPase活性定位变化与NaHCO₃胁迫下星星草抗盐碱能力的关系。结果表明: 在正常状态下, 根毛区细胞质内Ca²⁺较少, 主要位于质膜附近和液泡中, Ca²⁺-ATPase主要定位于质膜和液泡膜, 有一定活性。在0.448%NaHCO₃胁迫下, 根毛区细胞质中Ca²⁺增多, 液泡中Ca²⁺减少, 且主要集中于液泡膜附近, 质膜和液泡膜Ca²⁺-ATPase活性明显升高。在1.054%NaHCO₃胁迫下,细胞质中分布的Ca²⁺增多, 而液泡中Ca²⁺极少, Ca²⁺-ATPase活性也降低。以上结果表明, Ca²⁺亚细胞定位和Ca²⁺-ATPase活性变化在星星草响应NaHCO₃胁迫的信号传递过程中具有重要作用。     

植物细胞质膜H+-ATPase的调控

综述了质膜H ATPase在转录、翻译及翻译后水平上所受调控现象及其机制的研究进展     

pH-dependent regulation of the α-subunit of H+-K+-ATPase (HKα2)

The H(+)-K(+)-ATPase α-subunit (HKα(2)) participates importantly in systemic acid-base homeostasis and defends against metabolic acidosis. We have previously shown that HKα(2) plasma membrane expression is regulated by PKA (Codina J, Liu J, Bleyer AJ, Penn RB, DuBose TD Jr. J Am Soc Nephrol 17: 1833-1840, 2006) and in a separate study demonstrated that genetic ablation of the proton-sensing G(s)-coupled receptor GPR4 results in spontaneous metabolic acidosis (Sun X, Yang LV, Tiegs BC, Arend LJ, McGraw DW, Penn RB, Petrovic S. J Am Soc Nephrol 21: 1745-1755, 2010). In the present study, we investigated the ability of chronic acidosis and GPR4 to regulate HKα(2) expression in HEK-293 cells. Chronic acidosis was modeled in vitro by using multiple methods: reducing media pH by adjusting bicarbonate concentration, adding HCl, or by increasing the ambient concentration of CO(2). PKA activity and HKα(2) protein were monitored by immunoblot analysis, and HKα(2) mRNA, by real-time PCR. Chronic acidosis did not alter the expression of HKα(2) mRNA; however, PKA activity and HKα(2) protein abundance increased when media pH decreased from 7.4 to 6.8. Furthermore, this increase was independent of the method used to create chronic acidosis. Heterologous expression of GPR4 was sufficient to increase both basal and acid-stimulated PKA activity and similarly increase basal and acid-stimulated HKα(2) expression. Collectively, these results suggest that chronic acidosis and GPR4 increase HKα(2) protein by increasing PKA activity without altering HKα(2) mRNA abundance, implicating a regulatory role of pH-activated GPR4 in homeostatic regulation of HKα(2) and acid-base balance.     

Is the Ca2+-ATPase from sarcoplasmic reticulum also a heat pump?

We calculate, using the first law of thermodynamics, the membrane heat fluxes during active transport of Ca²⁺ in the Ca²⁺-ATPase in leaky and intact vesicles, during ATP hydrolysis or synthesis conditions. The results show that the vesicle interior may cool down during hydrolysis and Ca²⁺-uptake, and heat up during ATP synthesis and Ca²⁺-efflux. The heat flux varies with the SERCA isoform. Electroneutral processes and rapid equilibration of water were assumed. The results are consistent with the second law of thermodynamics for the overall processes. The expression for the heat flux and experimental data, show that important contributions come from the enthalpy of hydrolysis for the medium in question, and from proton transport between the vesicle interior and exterior. The analysis give quantitative support to earlier proposals that certain, but not all, Ca²⁺-ATPases, not only act as Ca²⁺-pumps, but also as heat pumps. It can thus help explain why SERCA 1 type enzymes dominate in tissues where thermal regulation is important, while SERCA 2 type enzymes, with their lower activity and better ability to use the energy from the reaction to pump ions, dominate in tissues where this is not an issue.

Evidence for the involvement of (Cu-ATP)2− in the inhibition of human erythrocyte (Ca2+ + Mg2+-ATPase by copper

The effects of copper on the activity of erythrocyte $\text{(}\text{Ca}{}^{\mathrm{2+}}\text{+}\text{Mg}{}^{\mathrm{2+}}\text{)-}\text{ATPase}$ have been tested on membranes stripped of endogenous calmodulin or recombined with purified calmodulin. The interactions of copper with Ca²⁺, calmodulin and (Mg-ATP)^2? were determined by kinetic studies. The most striking result is the potent competitive inhibition exerted by (Cu-ATP)^2? against (Mg-ATP)^2?$\text{K}{}_{\mathrm{<mtext>i</mtext>}}\text{= 2.8 μ}\text{M}$), while free copper gives no characteristic inhibition. Our results also demonstrate that copper does not compete with calcium either on the enzyme or on calmodulin. The fixation of calmodulin on the enzyme is not altered in the presence of copper as shown by the fact that the dissociation constant remains unaffected. It may be speculated that (Cu-ATP)^2? is the active form of copper, which could plausibly be at the origin of some of the pathological features of erythrocytes observed in conditions associated with excess copper.     

草鱼和鲢鱼肌球蛋白Ca2+-ATPase热力学活化参数的季节变化

肌球蛋白构成了鱼肌肉肌原纤维蛋白的50%以上,它的性质主要决定了肌原纤维蛋白的性质,进而影响了鱼肌肉蛋白质的加工适性.本文对来自春季(4月份)、夏季(8月份)、秋季(11月份)和冬季(1月份)的草鱼(Ctenopharyngodon idellus Valenciennes)和鲢鱼(Hypophthalmichthys molitrix Valenciennes)骨骼肌肌球蛋白进行提取和纯化,通过测定不同季节的两种淡水鱼肌球蛋白Ca2 -ATPase的热力学活化参数,从化学反应的热力学理论上证明了淡水鱼肌球蛋白的热稳定性与季节温度变化之间的关系.研究结果表明,草鱼和鲢鱼肌球蛋白Ca2 -ATPase的热力学活化参数活化能Ea、活化焓ΔH 和活化熵ΔS 伴随栖息环境温度的上升而增大,而活化自由能ΔG 的变化幅度很小,证明了肌球蛋白的热稳定性夏季鱼明显优于冬季鱼,而春季鱼和秋季鱼分别与夏季鱼和冬季鱼的相似.另一方面,ΔH 与ΔS 之间显示了良好的线性相关,证明了淡水鱼能够通过其肌肉蛋白质反应的活化焓与活化熵之间的互补效应,以应对自然界季节环境温度的变化.     

Calmodulin antagonizes amyloid-β peptides-mediated inhibition of brain plasma membrane Ca2+-ATPase

The synaptosomal plasma membrane Ca²⁺-ATPase (PMCA) plays an essential role in regulating intracellular Ca²⁺ concentration in brain. We have recently found that PMCA is the only Ca²⁺ pump in brain which is inhibited by amyloid-β peptide (Aβ), a neurotoxic peptide implicated in the pathology of Alzheimer's disease (AD) [1], but the mechanism of inhibition is lacking. In the present study we have characterized the inhibition of PMCA by Aβ. Results from kinetic assays indicate that Aβ aggregates are more potent inhibitors of PMCA activity than monomers. The inhibitory effect of Aβ could be blocked by pretreating the purified protein with Ca²⁺-calmodulin, the main endogenous activator of PMCA, and the activity of truncated PMCA lacking the calmodulin binding domain was not affected by Aβ. Dot-overlay experiments indicated a physical association of Aβ with PMCA and also with calmodulin. Thus, calmodulin could protect PMCA from inhibition by Aβ by burying exposed sites on PMCA, making them inaccessible to Aβ, and also by direct binding to the peptide. These results suggest a protective role of calmodulin against neuronal Ca²⁺ dysregulation by PMCA inhibition induced by Aβ.