黄瓜叶绿素降解关键酶基因CsPAO的克隆与分析

该研究以黄瓜“津春2号”cDNA为模板,采用RT PCR方法克隆得到黄瓜叶绿素降解关键酶(pheophorbide a oxygenase,PAO)基因(CsPAO),对其进行亚细胞定位观察,并采用实时荧光定量PCR技术和生物信息学技术,分析了CsPAO基因的表达模式及其编码蛋白的特性。结果表明：(1)CsPAO编码545个氨基酸,理论等电点为6.09,蛋白相对分子质量为61.02 kD。蛋白预测发现,黄瓜CsPAO属于不稳定蛋白,具有2个蛋白结合位点,且存在跨膜现象。(2)荧光定量PCR结果表明,CsPAO基因表达响应水杨酸(SA)、茉莉酸 (JA)和赤霉素(GA₃)的调控,在高温(42 ℃)和低温(4 ℃)处理下CsPAO基因的表达量显著上升并达到最高,但黑暗处理对CsPAO基因表达没有影响;在黄瓜不同组织中花的表达显著高于根、茎、叶、萼、须、果。(3)亚细胞定位结果表明,CsPAO蛋白定位于叶绿体内。(4)系统进化树分析显示,黄瓜CsPAO与葫芦科植物苦瓜、西葫芦、南瓜、笋瓜等亲缘关系较近。本研究结果为进一步揭示黄瓜叶绿素降解的分子机制奠定了基础。     

脱镁叶绿素a-光动力疗法抗前列腺癌的研究进展

脱镁叶绿素a是一种从高等植物和海藻中提取的化合物,具有抗菌、抗炎、抗病毒、抗氧化和抗肿瘤的药理作用;作为一种天然光敏剂,其与光动力疗法联用,对于多种肿瘤表现出良好的抑制效果.近年来,脱镁叶绿素a在癌症治疗方面的研究取得了重大进展,在抗前列腺癌方面表现出巨大的潜力.该文从脱镁叶绿素a结构特征、应用及抗前列腺癌作用机制方面...     

植物脱镁叶绿素替代RS601反应中心中的细菌脱镁叶绿素的研究

在一定的温度和丙酮的协同作用下,植物脱镁叶绿素a可置换紫细菌RS601光合反应中心的细菌脱镁叶绿素而形成含有植物脱镁叶绿素的紫细菌光合反应中心(简称Phe a RC).当协同作用15和60min时,反应中心的细菌脱镁叶绿素分别被替代了50%和71%.在Phe a RC中,细菌脱镁叶绿素的QX(537nm)和QY(758nm)特征峰显著下降,而出现植物脱镁叶绿素的QX(509/542nm)和QY(674nm)特征峰.排除温度和丙酮的影响,替代时间为15或60min的Phe a RC的光化学活性分别为对照的78%或71%,其电化学特性也有所变化.     

大亚湾冬季沉积物中叶绿素含量与分布

2012年2月(冬季)对大亚湾海域沉积物叶绿素a (Chl-a)及其降解产物脱镁叶绿酸( Pha-a)的含量与分布进行了研究,并探讨了环境因子(水深、温度、透明度、粒度、营养盐、TOC)对表层沉积物Chl-a含量分布的影响.结果表明:Chl-a和Pha-a是同源的,随沉积物深度增加二者皆呈递减趋势,个别站位Chl-a和Pha-a出现许多小突跃,这可能与生物扰动和环境变化有关;表层(0～l cm)Chl-a和Pha-a的含量变化范围分别为0.98 ～ 1.55μg·g-1和3.97～6.65 μg·g-1,平均含量分别为1.24和5.55 μg ·g-1.Chl-a含量变化是各种环境因子共同作用的结果,其中最主要的影响因素是沉积物底质颗粒的大小,高比例砂含量的底质能为底栖藻类提供更多的生存空间,但不利于沉积物中Pha-a的保存;比较了沉积物与上层水体中平均Chl-a相对含量,并与其他海域相比较,发现该海域沉积物中Chl-a含量较高,与水体Chl-a含量比值为50.28％ ～ 150.79％,近海沉积物中底栖初级生产力不可忽视.     

久效磷对扁藻的损伤作用 Ⅱ. 叶绿素a降解与活性氧损伤的相关性

扁藻细胞在久效磷的毒性胁迫下,随着胁迫强度的增加,叶绿素a降解加剧,其含量逐渐降低。分析表明,叶绿素a含量的降低与活性氧O₂^-含量、膜脂过氧化产物丙二醛(MDA)含量及细胞的电解质外渗率呈显着负相关;而与细胞超氧化物歧化酶(SOD)活性及过氧化物酶(POD)活性的降低呈显着正相关,说明久效磷胁迫下扁藻细胞叶绿素a降解与其活性氧的损伤有明显相关性。     

久效磷对扁藻的损伤作用Ⅱ.叶绿素a降解与活性氧损伤的相关性

扁藻细胞在久效磷的毒性胁迫下,随着胁迫强度的增加,叶绿素ａ降解加剧,其含量逐渐降低．分析表明,叶绿素ａ含量的降低与活性氧Ｏ２－含量、膜脂过氧化产物丙二醛（ＭＤＡ）含量及细胞的电解质外渗率呈显著负相关;而与细胞超氧化物歧化酶（ＳＯＤ）活性及过氧化物酶（ＰＯＤ）活性的降低呈显著正相关,说明久效磷胁迫下扁藻细胞叶绿素ａ降解与其活性氧的损伤有明显相关性．     

植物叶绿素降解途径及其分子调控

文章介绍了近年来在叶绿素降解产物结构解析和关键酶基因克隆方面的最新成果,以及在此基础上的叶绿素降解途径修正及其分子调控机理研究。     

紫细菌光合反应中心中细菌脱镁叶绿素被置换后的光 …

采用新型表面活性剂ＬＤＡＯ,结合ＤＥＡＥ－纤维素层析法,我们提纯了紫细菌６０１的光合反应中心,在一定温度和丙酮的协同作用下,外加的植物脱镁叶绿素ａ可取代反应中心细菌脱镁叶绿素,形成含有脱镁叶绿素ａ的紫细菌光合反应中心。     

植物叶绿素的降解

文章介绍近几年来植物叶绿素降解途径以及与其相关几种酶的研究进展。     

紫细菌光合反应中心中细菌脱镁叶绿素被置换后的光化学活性

采用新型表面活性剂ＬＤＡＯ,结合ＤＥＡＥ纤维素层析法,我们提纯了紫细菌Ｒｈｏｄｏｂａｃｔｅｒｓｐｈａｅｒｏｉｄｅｓ６０１的光合反应中心。在一定温度和丙酮的协同作用下,外加的植物脱镁叶绿素ａ可取代反应中心细菌脱镁叶绿素,形成含有脱镁叶绿素ａ的紫细菌光合反应中心（ＰｈｅａＲＣ）。当协同作用１５ｍｉｎ和６０ｍｉｎ时,反应中心中的细菌脱镁叶绿素分别被替代了５０％和７１％。在ＰｈｅａＲＣ中,细菌脱镁叶绿素的ＱＸ（５３７ｎｍ）和ＱＹ（７５８ｎｍ）特征峰显著下降,而出现高等植物脱镁叶绿素的ＱＸ（５０９／５４２ｎｍ）和ＱＹ（６７４ｎｍ）特征峰。排除温度和丙酮的影响,替代时间为１５ｍｉｎ或６０ｍｉｎ的ＰｈｅａＲＣ的光化学活性分别为对照的７８％或７１％。     

Chlorophyll b to chlorophyll a conversion precedes chlorophyll degradation in Hordeum vulgare L.

This study reveals by in vivo deuterium labeling that in higher plants chlorophyll (Chl) b is converted to Chl a before degradation. For this purpose, de-greening of excised green primary leaves of barley (Hordeum vulgare) was induced by permanent darkness in the presence of heavy water (80 atom % (2)H). The resulting Chl a catabolite in the plant extract was subjected to chemical degradation by chromic acid. 3-(2-Hydroxyethyl)-4-methyl-maleimide, the key fragment that originates from the Chl catabolite, was isolated. High resolution (1)H-, (2)H-NMR and mass spectroscopy unequivocally demonstrates that a fraction of this maleimide fragment consists of a mono-deuterated methyl group. These results suggest that Chl b is converted into Chl a before degradation. Quantification proves that the initial ratio of Chl a:Chl b in the green plant is preserved to about 60-70% in the catabolite composition isolated from yellowing leaves. The incorporation of only one deuterium atom indicates the involvement of two distinguishable redox enzymes during the conversion.     

Peroxidase-mediated chlorophyll degradation in horticultural crops

One of the symptoms of senescence in harvested horticultural crops is the loss of greenness that comes with the degradation of chlorophyll (Chl). With senescence, peroxidase, which is involved in Chl degradation, increased greatly in stored horticultural crops. C13²-hydroxychlorophyll a, an oxidized form of Chl a, is formed in vitro through Chl oxidation by peroxidase. Peroxidase mediates Chl degradation in the presence of phenolic compounds such as p-coumaric acid and apigenin, which have a hydroxyl group at the p-position. Apparently, not all phenolic compounds are able to degrade Chl in this system, and their effectiveness appears to depend on their molecular configuration. In peroxidase-mediated Chl degradation, peroxidase oxidizes the phenolic compounds with hydrogen peroxide and forms phenoxy radical; then, the phenoxy radical oxidizes Chl and its derivatives to colorless low molecular weight compounds through the formation of C13²-hydroxychlorophyll a,a fluorescent Chl catabolite and a bilirubin-like compound as an intermediate. In addition to the phenoxy radical, superoxide anion, which is formed in the peroxidase-catalyzed reaction, might be involved in Chl oxidation. Moreover, Chl degradation by peroxidase seems to occur in the chloroplast and/or the vacuole. The involvement of peroxidase in Chl degradation in senescing horticultural crops is also discussed.     

Enzymatic degradation of chlorophyll a by marine phytoplankton in vitro

The enzymatic degradation of chlorophyll a and the formation of chlorophyllide a, phaeophytin a, and phaeophorbide a were detected in vitro in several species of marine phytoplankton. Loss of phytol and Mg²⁺ were found to be catalysed by chlorophyllase and a magnesium-releasing enzyme, respectively. The activities of the two enzymes could be distinguished from each other by inhibiting with Mg²⁺ and/or p-chloromercurobenzoate. Both enzymes are activated by cell disintegration. Degradation products were not detected spectrophotometrically in vivo. Additionally, in some species, chlorophyll a was degraded to products which do not absorb visible light.     

Bacterial degradation of styrene involving a novel flavin adenine dinucleotide-dependent styrene monooxygenase

By using styrene as the sole source of carbon and energy in concentrations of 10 to 500 microM, 14 strains of aerobic bacteria and two strains of fungi were isolated from various soil and water samples. In cell extracts of 11 of the bacterial isolates, a novel flavin adenine dinucleotide-requiring styrene monooxygenase activity that oxidized styrene to styrene oxide (phenyl oxirane) was detected. In one bacterial strain (S5), styrene metabolism was studied in more detail. In addition to styrene monooxygenase, cell extracts from strain S5 contained styrene oxide isomerase and phenylacetaldehyde dehydrogenase activities. A pathway for styrene degradation via styrene oxide and phenylacetaldehyde to phenylacetic acid is proposed.     

Pheophorbide a Content and Chlorophyllase Activity in Green Tea

We investigated the total content of pheophorbide a (PB a), which is sum of the contents of newly produced PB a, including PB a initially present and that converted from chlorophyllide a (Chd a) by the chlorophyllase reaction during incubation, in green tea samples, and found that the total content of PB a markedly increased in both Sencha and Matcha, compared with the initially present PB a content in each. This result demonstrates that chlorophyllase activity still remains in green tea, even after processing fresh green leaves. A comparison of the total contents of PB a produced during the incubation of chlorophyll a (Chl a) with Sencha and fresh green leaf acetone powder indicates that the ratio of chlorophyllase activity in Sencha and in fresh green leaves was about 1:20.     

Molecular cloning and characterization of a chlorophyll degradation regulatory gene from bamboo

Leaf senescence constituted the final stage of leaf development and it is always accompanied by the leaf yellowing. The non-yellowing gene (NYE1), initially identified from Arabidopsis in our laboratory, is a key regulatory gene responsible for chlorophyll degradation during senescence. In this study, an orthologue of AtNYE1 was isolated from the bamboo (Bambusa emeiensis cv. Viridiflavus) and tentatively named BeNYE1. The full length sequence of 1 386 bp contains an open reading frame of 801 bp. The protein encoded by BeNYE1 consists of 266 amino acids. Sequence analysis revealed that BeNYE1 had high similarity with other NYE/SGR proteins from various monocotyledon species. BeNYE1 was strongly induced by natural senescence and dark-induced senescence in bamboo. Driven by a 1.5 kb upstream fragment of AtNYE1, BeNYE1 could rescue the stay-green phenotype of nye1-1. The constitutive over-expression of BeNYE1 could accelerate the chlorophyll degradation. These results indicated that BeNYE1 might play an important role in the regulation of chlorophyll degradation during leaf senescence in bamboo.     

Bacterial degradation of styrene involving a novel flavin adenine dinucleotide-dependent styrene monooxygenase.

By using styrene as the sole source of carbon and energy in concentrations of 10 to 500 microM, 14 strains of aerobic bacteria and two strains of fungi were isolated from various soil and water samples. In cell extracts of 11 of the bacterial isolates, a novel flavin adenine dinucleotide-requiring styrene monooxygenase activity that oxidized styrene to styrene oxide (phenyl oxirane) was detected. In one bacterial strain (S5), styrene metabolism was studied in more detail. In addition to styrene monooxygenase, cell extracts from strain S5 contained styrene oxide isomerase and phenylacetaldehyde dehydrogenase activities. A pathway for styrene degradation via styrene oxide and phenylacetaldehyde to phenylacetic acid is proposed.