叶绿素缺乏大麦突变体光系统II色素蛋白复合物的研究

同野生型大麦相比,突变大麦光系统Ⅱ捕光色素蛋白复合物的含量明显降低,其多肽组分也发生了变化;２６ｋＤ的多肽缺失,２４ｋＤ、２７ｋＤ和３０ｋＤ多肽含量减少。ＲＮＡ印迹杂交结果表明突变大麦中ｃａｂ基因的表达与野生大麦基本一致。说明突变大麦中ＬＨＣＩＩ多肽的缺乏不是在转录水平引起的障碍,而很可能是受转录后水平的调节。突变大麦叶绿体的内膜系统处于发育的初级阶段,基粒较少,类囊体膜的垛叠也受到很大的限制,这     

叶绿素缺乏的大麦突变体的光合作用和叶绿素荧光

和早熟３号大麦（野生型）（ＤＧ）相比,黄化大麦（突变体）（ＬＧ）叶片Ｃｈｌ含量低,而Ｃｈｌ ａ／ｂ较高;光合速率低,表观量子效率低,而饱和光强较高,气孔导度较高;荧光参数Ｆｏ低,而ＰＳⅡ的光化学效率（Ｆｖ／Ｆｍ）以及Ｔ１／２、Ｆｍ／Ｆｏ、Φｅ均高。以单位叶绿素计算,黄化大麦的ＰＳⅡ电子传递活性和全链电子传递活性较高,而ＰＳＩ电子传递活性较低。推测黄化大麦ＰＳⅡ的光化学效率增高可能是由于其ＰＳⅡ向Ｐ     

叶绿素缺乏大麦突变体光系统I色素蛋白复合物的研究

同野生型大麦相比,突变大麦光系统ＩＩ捕光色素蛋白复合物的含量明显降低,其多肽组分也发生了变化：２６ｋＤ的多肽缺失,２４ｋＤ、２７ｋＤ和３０ｋＤ多肽含量减少。ＲＮＡ印迹杂交结果表明突变大麦中ｃａｂ基因的表达与野生大麦基本一致,说明突变大麦中ＬＨＣＩ多肽的缺乏不是在转录水平引起的障碍,而很可能是受转录后水平的调节。突变大麦叶绿体的内膜系统处于发育的初级阶段,基粒较少,类囊体膜的垛叠也受到很大的限制,这可能与２６ｋＤ多肽的缺失有关。     

叶绿素缺乏的大麦突变体的光合作用和叶绿素荧光

和早熟３号大麦（野生型）（ＤＧ）相比,黄化大麦（突变体）（ＬＧ）叶片Ｃｈｌ含量低,而Ｃｈｌａ／ｂ较高;光合速率低,表现量子效率低,而饱和光强较高,气孔导度较高;荧光参数Ｆｏ低,而ＰＳⅡ的光化学效率（Ｆｖ／Ｆｍ）以及Ｔ１／２、Ｆｍ／Ｆｏ、Φｅ均高。以单位叶绿素计算,黄化大麦的ＰＳⅡ电子传递活性和全链电子传递活性较高,而ＰＳⅠ电子传递活性较低。推测黄化大麦ＰＳⅡ的光化学效率增高可能是由于其ＰＳⅡ向ＰＳⅠ传递的激发能较少,是对Ｃｈｌ含量低的一种补偿。     

蓝藻叶绿素蛋白复合体的分离研究

蓝藻类囊体膜用声波超时处理,然后在4℃下用低浓度的LDS增溶,并经改进的SDS-聚丙烯酰胺凝胶电泳后被分离成15条绿色的带. 其中CPa1～CPa6有着相似的吸收光谱. 这6个组分的低温荧光光谱也很相似,其荧光发射光谱的发射峰都位于685 nm处,表明它们都属于光系统Ⅱ叶绿素a蛋白复合体. 该系统对光系统Ⅱ的分离能力是传统电泳的3倍.     

叶绿素缺乏的大麦突变体PSⅡ光化学效率较高的原因(英)

在相同的叶绿素浓度下,叶绿素缺乏的大麦突变体的叶绿体在450～480nm和600～640nm波长范围的光吸收值比野生型略高,而在400～440nm和700～740nm波长范围的光吸收值明显高于野生型;突变体叶片和叶绿体的低温（77K）荧光发射强度较低,而两个光系低温荧光产量的比值（r685／F735）较高;失去Mg2 后,突变体和野生型的F685／F735和Fv／Fm均降低,但突变体的降低幅度较小;突变体的叶绿体中基粒片层数目较少、长度较短,而间质片层较长。这些结果表明,大麦突变体PSⅡ向PSⅠ的激发能转移较少是其PSⅡ光化学效率较高的重要原因。     

光下叶绿素缺乏的大麦突变体NYB中叶绿素合成再探

研究一种原叶绿素酯还原酶（POR）的大麦突变体Ⅳ阳光下合成叶绿素的结果表明,NYB中PORB的含量比野生型低。NYB前质体中原片层体的大小和数量与野生型差不多,但其结构比野生型的松散。暗中生长的突变体内POR蛋白复合物LHPP比野生型少。不同光照强度下叶绿素积累的结果显示,光照度越强,突变体与野生型的叶绿素差异越显著。由于porB是单拷贝的,所以推测突变体中部分porB mRNA可能产生错误的剪切拼接,以致光下突变体Ⅳ阳仍然能合成叶绿素。     

杂交小麦不同发育时期叶绿素蛋白复合体的变化

杂交小麦“９０１”在不同发育时期及不同叶位,叶片色素蛋白复合体含量均高于对照品种陕２２９,尤其表现在“９０１”的旗叶,而陕２２９的倒二叶在籽粒形成后期、灌浆期,略高于“９０１”,倒三叶在籽粒发育后期至成熟期“９０１”的含量要高于陕２２９。叶绿素蛋白复合体分析结果说明,在不同发育时期、不同叶位,“９０１”叶片色素蛋白复合体中含有较高的叶绿素b,尤其表现在籽粒形成后期的旗叶上。２个品种在不同发育时期,不同时叶位之间,色素蛋白复合体组分上没有差异。仅表现在成熟期,陕２２９的倒三叶各色素带含量减少,并且消失１条叶绿素蛋白复合体带,初步认为此带是ＬＨＣＰ的寡聚体之一,而“９０１”比陕２２９晚７d倒二叶上表现出样的现象,表明“９０１”叶片色素蛋白复合体抗衰老能力很强。     

精胺对离体大麦叶片中叶绿素和蛋白质含量的影响

10~(-3)mol/L精胺(Spermine)能够有效地延缓离体大麦叶片的叶绿素、蛋白质总量的下降,并保持chla/chlb值,阻止非水溶性蛋白质的降解,提高结合态叶绿素相对含量。精胺对幼叶的作用大于其对老叶的作用。     

Chlorophyll-Protein Complexes of Blue-Green Algae Isolated by a New Gel Electrophoresis System with High Resolving Power

At least 13 chlorophyll bands from the thylakoid membranes of blue-green algae could be clearly resolved by SDS-PAGE employing a new improved procedure. They were designated as CPIa, CPIb, CPIc, CPId, CPIe, CPIf, CPIg, CHIh, CPal, CPa2, CPa3, CPa4 and FC. 8 chlorophyll-protein complexes, CPIa-CPIh, had the same absorption spectrum at 676 nm in the red and 436 nm in the blue region. They belonged to the chlorophyll-protein complexes of PS Ⅰ. 4 chlorophyll-protein complexes, CPal-CPa4, had a red absorption peak at 670­672 nm and a blue one at 436 nm. Their fluorescence emission peak at 77K was at 685 nm. They were chlorophyll-protein complexes of PS Ⅱ.     

一个高分辨率的凝胶电泳系统及其从蓝藻中分离的叶绿素蛋白复合体

用一高分辨率的凝胶电泳系统从蓝藻类囊体膜中分离出至少１３ 个清晰的叶绿素带,它们是ＣＰＩａ、ＣＰＩｂ、ＣＰＩｃ、ＣＰＩｄ、ＣＰＩｅ、ＣＰＩｆ、ＣＰＩｇ、ＣＰＩｈ、ＣＰａ１、ＣＰａ２、ＣＰａ３、ＣＰａ４ 和ＦＣ,其分辨率较传统方法高出１ 倍多。ＣＰＩａ—ＣＰＩｈ ８ 种组分有相同的吸收光谱,其红峰和蓝峰的位置分别位于６７６ ｎｍ 和４３６ ｎｍ 处。它们都属于光系统Ｉ叶绿素蛋白复合体。ＣＰａ１—ＣＰａ４ ４ 种组分的光谱性质亦基本相同,其吸收峰的位置分别位于６７０—６７２ ｎｍ 和４３６ ｎｍ 处,而低温荧光发射峰的位置都位于６８５ ｎｍ 处。它们都属于光系统Ⅱ叶绿素蛋白复合体     

一种分析叶绿体类囊体膜色素蛋白复合物的蓝绿温和胶电泳系统

采用蓝绿温和胶电泳系统可以非常有效地分离叶绿体蛋白质复合物,包括PSⅠ, PSⅡ, ATP合酶,细胞色素b₆f复合物,捕光色素复合物和1,5-二磷酸核酮糖羧化酶.还结合SDS-聚丙烯酰胺凝胶电泳将叶绿体多亚基复合物的50多种蛋白质分开,利用免疫印迹对蛋白质复合物进行了初步鉴定,同时还应用蓝色温和胶电泳分析基质、基粒类囊体复合物的组成.     

The Chlorophyll-Protein Complexes Resolved from Ultrasonically Broken Thylakoid Memb-ranes of Blue-Green Algae

When the thylakoid membranes of blue-green algae were broken by ultrasonic vibrations and subjected to polyacrylamide gel electrophoresis at 4℃, six green zones were resolved. They were designated as CPIa, CPlb, CPI; CPal, CPa2, and FC. The absorption spectrum of CPI had a red maximum at 674 nm and a peak in the blue at 435 nm. It was identified as PS chlorophyll a-protein Complex, but was contaminated with minor PSⅡ which was implied by the appearance of fluorescence emission peak at 680 nm besides the main one at 725 nm at 77 K. The spectral properties of CPIa and CPlb were similar to that of CPl. The absorption spectra of CPa1 and CPa2 were similar, both having red maxima at 667 nm and peaks in the blue at 431.5 nm. Their fluorescence emission had the same peaks at 684 nm at 77 K indicating that they belonged to PSⅡ. It was recognized that CPal of 47 kD is the reaction center complex of photosystem Ⅱ and CPa2 of 40 kD is the internal antenna complex of photosystem Ⅱ. The spectral characteristics of the chlorophyll-protein complexes resolved by ultrasonic method were similar to those of the same complexes resolved by SDS solubilization, except the absorbance positions of CPa1 and CPa2 in the blue peak and the red one which shifted to blue about 3–5 nm. It was calculated that in thylakoid membranes of blue-green algae 40.93% chlorophyll was in PSⅠ, while 38.78% of chlorophyll in PSⅡ. The difference of chlorophyll contents between PSⅠ and PSⅡ was only 2.15%. Concerning the fact that minor PSⅡ compound remained in the part of PSⅠ zones, it might be concluded that the distribution of chlorophyll between PSⅠ and PSⅡ in blue-green algae was equal. This result was in agreement with the hypothesis that PSⅠ and PSⅡ operates in series in photosynthetic electron transport.     

大麦黄矮病毒运动蛋白是RNA沉默抑制因子

大麦黄矮病毒(barley yellow dwarf virus,BYDV)属黄症病毒科家族,其基因组包含6个开放阅读框(open reading frames,ORFs).将BYDV的6个基因分别克隆到pWEIMING101载体上,得到重组基因.电击转化农杆菌后,利用农杆菌瞬时表达方法渗透注射转GFP基因的本氏烟草16c植株的叶片,在长波长紫外灯下观察GFP的表达,并通过Northern blot证明所得现象.研究结果表明,BYDV的PAV株系ORF4编码的运动蛋白(movement protein,MP)是RNA沉默抑制因子,其表达可以抑制局部和系统RNA沉默.BYDV-MP与GFP的双链RNA(dsGFP)共表达后仍能抑制RNA沉默,荧光强度与叶片中GFP的mRNA和其沉默降解形成的siRNA的量有对应关系,其N端核定位序列对抑制局部基因沉默起主要作用,第5、6位氨基酸是抑制基因沉默的关键氨基酸.BYDV-MP单独渗透注射的部位均产生细胞死亡.     

Dimerization,Oligomerization, and Aggregation of Human Amyotrophic Lateral Sclerosis Copper/Zinc Superoxide Dismutase 1 Protein Mutant Forms in Live Cells

More than 100 copper/zinc superoxide dismutase 1 (SOD1) genetic mutations have been characterized. These mutations lead to the death of motor neurons in ALS. In its native form, the SOD1 protein is expressed as a homodimer in the cytosol. In vitro studies have shown that SOD1 mutations impair the dimerization kinetics of the protein, and in vivo studies have shown that SOD1 forms aggregates in patients with familial forms of ALS. In this study, we analyzed WT SOD1 and 9 mutant (mt) forms of the protein by non-invasive fluorescence techniques. Using microscopic techniques such as fluorescence resonance energy transfer, fluorescence complementation, image-based quantification, and fluorescence correlation spectroscopy, we studied SOD1 dimerization, oligomerization, and aggregation. Our results indicate that SOD1 mutations lead to an impairment in SOD1 dimerization and, subsequently, affect protein aggregation. We also show that SOD1 WT and mt proteins can dimerize. However, aggregates are predominantly composed of SOD1 mt proteins.     

Immunological characterization of chlorophyll a/b-binding proteins of barley thylakoids

Monoclonal antibodies have been raised against the light-harvesting chlorophyll a/b-binding proteins of photosystem I (LHCI) using a photosystem (PS) I preparation (PSI-200) wild-type from barley (Hordeum vulgare L. cv. Svaløf's Bonus) as the antigen. These antibodies cross-reacted with a minor light-harvesting chlorophyll a/b-protein of PSII (Chla/b-P1=CP29), but not with the major one, LHCII (=Chla/b-P2^**). Similarly, a monoclonal antibody to Chla/b-P1, elicited by a PSII preparation as the antigen, cross-reacted with LHCI, but not LHCII. This explains why an antigen consisting of LHCII, free of LHCI, but contaminated with Chla/b-P1, can elicit antibodies which cross-react with LHCI. Immunoblot assays showed that LHCI and Chla/b-P1 have at least two epitopes in common. Immunogold labelling of thin-sectioned wild-type thylakoids confirmed a preferential localisation of Chla/b-P1 in grana partition membranes and LHCI in stroma lamellae. The presence of LHCI was demonstrated in barley mutants lacking the PSI reaction centre (viridis-zb ⁶³) and chlorophyll b (chlorina-f2), and was correlated with the presence of long-wavelength (730 nm) fluorescence emission at 77 K. The mutant viridis-k ²³, which has a 77 K long-wavelength fluorescence peak at 720 nm, was shown by immune-blot assay to lack LHCI, although Chla/b-P1 was present.Abbreviations Chl-P chlorophyll-protein - CM Carlsberg Monoclonal - Da dalton - LHC light-harvesting complex - PAGE polyacrylamide gel electrophoresis - PSI, II photosystem I, II - PSI-200 PSI containing LHCI polypeptides - SDS sodium dodecyl sulphate     

The isolation and characterisation of a mutant of the C4 plant Amaranthus edulis deficient in NAD-malic enzyme activity

A mutant of Amaranthus edulis (Speg.) lacking activity of the C₄ leaf form of NAD-malic enzyme (ME; EC 1.1.1.39) has been isolated. Homozygous mutant (5% wild-type ME activity) and heterozygous (50% wild-type ME activity) F₂ plants were shown to contain both the α and β NAD-ME subunits in similar amounts to those detected in the wild-type leaves. The rate of photosynthetic CO₂ assimilation was reduced in the homozygous mutant to 5% of that observed for the wild-type leaves. Other C₄ enzymes were not down-regulated in the mutant plants. There was little difference in photosynthetic rate of the heterozygous plants compared to the wild-type, suggesting that NAD-ME exerts little control over the rate of C₄ photosynthesis, and that in the wild-type the enzyme has a very low control coefficient. The activity loss in the heterozygote may therefore be compensated by regulatory mechanisms that increase the activity of the enzyme in vivo. Data for bundle-sheath strands indicated that although the homozygous mutants were able to oxidise malate via the Krebs cycle, they were unable to convert malate to pyruvate and alanine via NAD-ME. Received: 2 April 1998 / Accepted: 7 May 1998