NADP—苹果酸酶活性变化及其在CAM运行中的调节

ＮＡＤＰ－苹果酸酶是ＣＡＭ植物一种重要脱羧酶。实验结果表明,专一ＣＡＭ植物瓦松和兼性ＣＡＭ植物长药景天及露花其ＮＡＤＰ－苹果酸酶活性昼高夜低;５－８月,兼性ＣＡＭ植物长药景天和露花随着Ｃ３光合型向ＣＡＤ型转化,其中ＮＡＤＰ－苹果酸活活性逐渐升高。     

CAM植物NADP─苹果酸的环境调节

专─ＣＡＭ植物瓦松（Ｏｒｏｓｔａｃｈｙｓｆｉｍｂｒｉａｔｕｓ）．兼性ＣＡＭ植物长药景天（Ｓｅｄｕｍｓｐｅｃｔａｂｉｌｅ、露花（Ｍｅｓｅｍｂｒｙａｎｔｈｅｍｕｍｃｏｒｄｉｆｏｌｉｕｍ）的ＮＡＤＰ─苹果酸酶活性在２５─５５℃的温度范围内,随着温度的升高而增强,高于５５℃后,活性下降;在ｐＨ６─９的范围内,反应系统ｐＨ７．５时,表现出最大活性。苹果酸是该酶反应底物,苹果酸浓度在１─５μｍｏｌ／Ｌ范围内,随着苹果酸浓度增加,酶活性上升,浓度高于５μｍｏｌ／Ｌ,对ＮＡＤＰ─苹果酸酶有抑制作用。光对该酶活性影响不大,在１３５０μｍｏｌ／ｍ２ｓ光照和黑暗条件下,处理４ｈ后,光下酶活性比黑暗下略有提高。看来,ＮＡＤＰ─苹果酸酶活性昼高夜低的变化规律并不是光／暗变化所致。     

苹果酸酶基因转化酿酒酵母的研究进展

微生物降酸是现代葡萄酒酿造工艺中重要环节之一。利用现代生物技术将粟酒裂殖酵母中的苹果酸酶基因和苹果酸通透酶基因共同转化到酿酒酵母中,构建苹果酸-酒精酵母,使之既能进行酒精发酵,又能分解苹果酸。主要对近些年粟酒裂殖酵母苹果酸酶性质、基因结构及其转化酿酒酵母的研究做了回顾与总结,并指出了有待于解决的问题。     

屋顶绿化植物佛甲草对温度梯度的生理生态适应

以屋顶生长的佛甲草为材料,通过光照培养箱进行不同温度条件处理,分别测量了叶片的CO2交换、叶绿素含量、叶绿素荧光参数以及植株不同部位的碳同位素比率变化(δ13C)。结果表明:持续高温/低温、较大的昼夜温差和叶表面风力的条件下,佛甲草为适应环境变化,光合会由C3代谢途径转变成景天酸代谢途径(CAM),是兼性CAM植物。短期降温会使叶片光系统Ⅱ(PSⅡ)发生不可逆失活,光合能力下降;复水后有助于PSⅡ的恢复和重建,而干旱天气会减缓恢复过程;在不利温度环境中生长的佛甲草老叶掉落较多,剩余叶片的叶绿素含量和Fv/Fm值增高,光合能力提高。δ13C测定结果显示,高温使嫩叶气孔导度降低,对成熟叶片气孔导度影响小,佛甲草茎杆虽然含有叶绿素,但没有明显的光合作用。     

卷枝毛霉中苹果酸酶同工酶V的酶学性质

【目的】探讨卷枝毛霉中苹果酸酶同工酶V的性质。【方法】克隆卷枝毛霉中编码苹果酸酶同工酶V的mel基因并在大肠杆菌BL21(DE3)中表达,利用His标签纯化获得了高纯度的重组酶BLME1,并进行酶学性质分析。【结果】该重组酶最适pH为8.0,最适温度为33℃,在此条件下酶活达到92.8 U/mg,对底物L-苹果酸和NADP~+的米氏常数K_m值为0.74960±0.06129 mmol/L和0.22070±0.01810 mmol/L,最大反应速度V_(max)分别为72.820±1.077 U/mg和86.110±1.665 U/mg。金属离子Mg~(2+)、Mn~(2+)、Co~(2+)、Ni~(2+)可以激活BLME1的活性,而Ca~(2+)、Cu~(2+)对BLME1活性则有抑制作用,中间代谢产物草酰乙酸和α-酮戊二酸也会抑制BLME1的活性,但琥珀酸却对BLME1有激活作用。【结论】本实验调查了卷枝毛霉苹果酸酶同工酶V的最适反应温度和pH、动力学参数,以及各种金属离子和中间代谢产物对酶活力的影响,这为以后深入研究该苹果酸酶的功能提供了理论依据和参考。     

苹果酸酶1及其对疾病调控的研究进展

苹果酸酶1(malic enzyme 1,ME1)是调节苹果酸代谢的关键酶,主要的生物学功能是维持细胞内氧化还原稳态、调节细胞能量代谢和合成生物分子,可影响细胞生长、分化、增殖和衰老等重要生命活动。近年来的研究表明,ME1与多种疾病的发生发展密切相关,且目前作为多种疾病潜在的治疗靶点备受关注。因此,本文将针对ME1的结构、生物学功能和转录调控机制以及与疾病的关系进行综述。     

山杏苹果酸酶基因的克隆与生物信息学分析

三羧酸转运体系(柠檬酸,苹果酸和丙酮酸)可为脂肪酸生物合成提供原料乙酰辅酶A与还原力NADPH;而苹果酸酶可催化苹果酸发生氧化脱羧而产生NADPH,是调控脂肪酸代谢的重要酶。山杏种子脂肪酸含量丰富、产油率高,是研究植物油脂代谢的良好材料。本研究以山杏为试材,采用RACE克隆和电子拼接相结合的方法,首次从山杏种子中克隆得到苹果酸酶基因(命名为SaME,GenBank上登记号为JX262381),该基因的cDNA编码区全长1923bp、编码640个氨基酸,预测其编码蛋白分子量为70.15kD、等电点为6.38;同时,运用生物信息学方法对SaME基因编码蛋白的理化特性、结构域和亚细胞定位等方面进行预测分析,结果显示,该蛋白定位在细胞的质体膜上,具有苹果酸酶活性,属于NADP-ME超家族。本文对山杏SaME基因的克隆与分析,为进一步探究苹果酸酶对油脂代谢的调控机制奠定了基础。     

大肠杆菌K12苹果酸酶的克隆、表达与纯化

以大肠杆菌K12基因组DNA为模板,PCR扩增得到NAD 依赖型苹果酸酶(NAD-ME)的全长基因,并克隆到载体pET24b( )中,得到表达质粒pET24b-ME。在IPTG诱导下,携带pET24b-ME的大肠杆菌BL21(DE3)高效表达分子量约为65 kDa的可溶性蛋白。重组NAD-ME经镍亲和层析纯化,比活达到100 U/mg以上。以上结果为深入研究苹果酸酶生物催化特性及其与辅酶的相互作用奠定了基础。     

土三七(Sedum aizoon)光合作用的季节转变和CAM的诱导

土三七的CO_3交换曲线在六月中旬以前呈C_3型的“方波”形,白天能大量吸收CO_2,夜间有CO_2放出,含酸量无显著的昼夜波动。夏季的其余时间,夜间有净的CO_2吸收(阶段Ⅰ),早晨有CO_2吸收峰(阶段Ⅱ),随后有光下的脱羧作用(阶段Ⅲ),并有明显的昼夜酸波动,呈CAM型,但无阶段Ⅳ。CAM季节出现前,水分胁迫可使成年叶子转变为CAM型,所需时间2～6天,再经浇灌(5 h)可恢复到原来的C_3型,表明诱导的CAM是可逆的。     

Behavior Study of NAD-Malic Enzyme in Facultative CAM Plants

Primary study of NAD-malic enzyme was carried out in facultative CAM plants Sedum aizoon and Mesembryanthemum cordifolium. When 5 mmol/L MnC12, 6 mmol/L Mal and 50μmol/L CoA are present, the optimum pH is 7. The activity of this enzyme fluctuates with the seasons, in July, its activity reaches the summit. Its activity has a diurnal change also, being higher during the day and lower during the night. After water stress, the activity of this enzyme increases 2—3 times. The authors consider that the CAM activity may be regulated by carboxylate and decarboxylate system together.     

兼性景天酸代谢植物CO2气体交换模式的环境调节

长药景天(Sedum speciubile)、土三七(S aizoon)及露花(Mesembryanihemum cord-ifolium)等三种兼性景天酸代谢(CAM)植物的CAM型植株,在阴天能全天吸收CO_2,无第Ⅲ阶段,与其在晴天的CO_2交换模式明显不同:而大叶伽蓝菜(Kalanchoe daigremontiana)、瓦松(Orostachys fimbriatus)及落地生根(Bryophyllum pinnatum)等三种专一性CAM植物,在阴天的气体交换模式仍有第Ⅲ阶段。说明在阴天能全天持续吸收CO_2的气体交换模式,只有兼性CAM植物具有。兼性CAM植物在阴天和晴天的气体交换模式间出现的差异,温度起着主导的作用。高温可加速苹果酸脱羧,温度对气体交换模式的影响即主要是通过影响脱羧速率实现的。专一性CAM植物由于晚上积累的苹果酸较多,使它们在阴天气温较低的条件下脱羧也较快,这是专一性CAM植物在阴天也不能全天吸收CO_2的重要原因。     

Photosynthesis but not CAM responded flexibly to changes in irradiance in Plectranthus marrubioides (Lamiaceae)

Well-watered plants of Plectranthus marrubioides Benth., a crassulacean acid metabolism (CAM) species naturally inhabiting sun exposed succulent places, were grown at photosynthetically active photon flux densities (PPFD) of either 150 (LL) or 300 (HL) μmol m-2 s-1 in a controlled environment. Photosynthesis of LL plants was saturated by irradiance of ca. 500 μmol m-2 s-1 while in HL plants saturation was not reached up to 1200 μmol m-2 s-1 and photosynthetic capacity was nearly 50 % higher than in the LL plants. However, maximum photon yield was 55 % lower and compensation irradiance was 25 % higher in LL plants. The former also had larger, more succulent leaves, i.e., they were morphologically more sun adapted. On the other hand, nocturnal accumulation of malic and citric acid, nighttime CO2 gain, and the low relative carbon recycling were independent of the prevailing PPFD. Furthermore, photosynthetic performance was flexibly and reversibly adjusted in HL plants after transfer to 600 or 150 μmol m-2 s-1 while nocturnal CO2 uptake was not influenced. Photosynthesis showed a high acclimation potential to high PPFD and patterns of gas exchange became more C3-like the higher the irradiance was, without a direct effect on CAM in P. marrubioides. This revised version was published online in June 2006 with corrections to the Cover Date.     

植物基因表达的代谢调控

通过通过代谢物,激素和环境因素对植物几种基因表达的影响,就植物基因表达的代谢调控进行了讨论,提出了开展有关这方面研究的一些设想。     

Effects of abscisic acid on CAM in Portulacaria afra

Water stress induces Crassulacean acid metabolism (CAM) in Portulacaria afra as manifested by day stomatal closure, organic acid fluctuation, and night CO₂ uptake. We now have evidence that abscisic acid treatment of leaves causes partial stomatal closure that is accompanied by the induction of CAM in a manner similar to water stress. There appears to be an inverse relationship between exogenous CO₂ uptake and decarboxylation of organic acids in that organic acids remain high during the day providing stomata are open. When stomata close, there is consumption of organic acids by decarboxylation. The hypothesis is that stomatal opening controls CAM in this species.This material is based upon work supported by the Science and Education Administration of the USDA under Competitive Grant No. 5901-0410-8-0018-0.     

Relationship between fern development and CAM in Pyrrosia piloselloides (L.) Price

Pyrrosia piloselloides (L.) Price is a constitutive CAM plant in the sporophytic phase of its life-cycle. Newly developed sporophytes, still attached to the gametophytes, showed signs of CAM expression in terms of diurnal changes in titratable acidity of the tissues. The gametophytes did not exhibit CAM. This revised version was published online in June 2006 with corrections to the Cover Date.     

CAM variations in the leaf-succulent Delosperma tradescantioides (Mesembryanthemaceae), native to southern Africa

Drought responses of diurnal gas exchange, malic acid accumulation and water status were examined in Delosperma tradescantioides , a succulent that grows in drought-prone microenvironments in summer rainfall and all-year rainfall regions of southern Africa. When well-watered, this species exhibited Crassulacean acid metabolism (CAM)-cycling, but its carbon fixation pattern changed during the development of drought, shifting to either low-level CAM or to CAM-idling. The rate and pattern of this change depended on environmental conditions, duration of water stress and leaf age. At the onset of drought, diurnal malate fluctuation increased, but was strongly depressed (by ca 70%) as drought continued, and when leaf water content and water potential were low (ca 35 and 50% of the initial levels, respectively). When rewatered, rates of growth and photosynthesis, gas exchange and water status recovered fully to pre-stressed values within two days. Whole-shoot carbon uptake rates suggested that leaf growth had continued unabated during a short-term (∼ one week) drought. This emphasises that CAM-idling allows the maintenance of active metabolism with negligible gas exchange when soil water is limiting. It is possible that old or senescent leaves may provide water for the expansion of developing leaves during initial periods of drought. Regardless of the water regime and environmental conditions, leaf nocturnal malate accumulation and water content were positively correlated and increased with leaf age. Thus the gradual loss of water from older mature leaves may induce CAM-idling, which reduces water loss. An important ecological consequence of this combination of CAM modes is the potential to switch rapidly between fast growth via C₃ gas exchanges when well-watered to water-conserving CAM-idling during drought.