铜绿假单胞菌铁摄取调节子Fur诱导表达突变株构建及表型分析

铁摄取调节子 (Ferric uptake regulator,Fur) 是细菌控制细胞内铁平衡的一类重要的调节子。铜绿假单胞菌Pseudomonas aeruginosa的fur为必需基因,不能直接敲除。文中通过构建诱导型缺失突变株Δfur/attB::PBAD-fur,来研究该基因对铜绿假单胞菌的生长、生物被膜形成、运动能力和抗氧应激能力等方面的影响。结果表明,当Fur低表达时,铜绿假单胞菌在高铁和低铁环境中出现了生长阻滞的现象;低表达Fur的铜绿假单胞菌抵抗H2O2的能力降低,形成生物被膜的能力减弱,游动、颤动 (Twitching) 和丛集 (Swarming) 运动能力也出现了减弱的现象。Fur的表达直接影响铜绿假单胞菌荧光嗜铁素的产量。在铜绿假单胞菌体内表达来自格瑞菲斯瓦尔德磁螺菌Fur超级家族中的蛋白,可以部分恢复铜绿假单胞菌荧光嗜铁素的产量。由此说明,Fur对铜绿假单胞菌的生长、生物被膜形成、抗氧应激能力和运动能力方面都起着至关重要的作用。本研究为铜绿假单胞菌的防治提供理论指导。     

wbnH2基因对北京欧文氏菌生物学特性及致病力的影响

【背景】北京欧文氏菌(Erwinia beijingensis)引起的刺芹侧耳细菌性软腐病(bacterial soft rot)给企业带来了严重的经济损失。wbnH2糖基转移酶基因在北京欧文氏菌中的生物学功能尚不明确。【目的】构建wbnH2基因的缺失株Δ-wbnH2和回补株C-wbnH2,探究wbnH2基因对北京欧文氏菌致病性的影响。【方法】采用同源重组方法构建北京欧文氏菌LMG 27579^TwbnH2基因缺失突变株Δ-wbnH2,并对基因缺失菌株的致病性、生长速度、运动能力、生物膜形成能力、黏附能力等生物学特性与野生型菌株进行比较分析;采用广宿主质粒pBBR1MCS2构建回补株C-wbnH2,排除了极性效应引起的突变株表型变化。【结果】与野生型菌株相比,基因缺失株Δ-wbnH2的生长速度无明显差别。但是wbnH2基因的缺失导致多糖分泌、生物膜形成能力、黏附能力、致病能力明显下降。【结论】wbnH2基因影响北京欧文氏菌多糖分泌、生物膜的形成能力、黏附能力及致病力,表明该基因在北京欧文氏菌致病过程中起重要作用,本研究为软腐病的防控提供了理论基础。     

细菌群体感应参与铜绿假单胞菌胞内聚羟基脂肪酸酯合成的调控

群体感应是细菌根据细胞密度变化调控基因表达的一种调节机制。铜绿假单胞菌中QS系统由lasI和rhlI合成的信号分子3OC12-HSL和C4-HSL以及各自的受体蛋白LasR、RhlR组成,它们以级联方式调控多个基因表达。【目的】研究细菌群体感应(QS)对聚羟基脂肪酸酯合成的调控。【方法】利用铜绿假单胞菌PAO1及其QS突变株为材料通过气相色谱、荧光定量PCR在生理和分子水平上研究QS对聚羟基脂肪酸酯合成的调控。【结果】QS信号分子合成抑制剂阿奇霉素处理铜绿假单胞菌PAO1和QS突变株导致胞内PHA积累量显著减少;铜绿假单胞菌PAO1中C4-HSL合成酶基因rhlI缺失突变株PAO210胞内PHA积累量与野生型无差别;而3OC12-HSL合成酶基因lasI缺失突变株PAO55、3OC12-HSL受体合成酶基因lasR缺失突变株PAO56以及lasI/lasR双缺失突变株PAO57胞内PHA含量与野生型相比明显减少;lasI和lasR的突变株体内PHA合成酶基因phaC1的表达量显著降低,信号分子3OC12-HSL回补实验使phaC1的表达量可恢复到野生株水平,但只可部分恢复lasI缺失导致的胞内PHA合成。【结论】由此推测,铜绿假单胞菌群体感应系统中lasI/lasR系统参与胞内聚羟基脂肪酸酯合成的调控。     

低温下阻遏蛋白MogR对单增李斯特菌鞭毛形成的影响

【目的】单核细胞增生李斯特菌(简称单增李斯特菌) (Listeria monocytogenes, Lm)是食源性病原菌,可以引发李斯特菌病(listeriosis)。Lm能在低温下生长,对冷藏食品的安全构成严重威胁,并对人类健康造成潜在的危害。Lm能低温生长与抑制鞭毛基因表达以减少鞭毛的合成有关。MogR是Lm鞭毛基因转录的阻遏蛋白,在机体里或37 ℃环境下具有阻遏作用,Lm不产生鞭毛;然而,当Lm处于20-0 ℃时MogR无阻遏作用,Lm产生鞭毛。我们研究发现在4 ℃生长条件下Lm鞭毛合成减少,但具体的分子机制尚未明确。本文探究在4 ℃下Lm鞭毛合成少与MogR起阻遏作用的关系。【方法】以Lm ATCC 19115为亲本株,分别构建了MogR和鞭毛丝蛋白FlaA的缺失株ΔmogR和ΔflaA (作为无鞭毛对照株)及其回补株cΔmogR和cΔflaA,测定了菌株在4、28、37 ℃时的运动性、鞭毛产生情况和鞭毛基因表达量,并对所构建的菌株进行了4、28、37 ℃时生长曲线的测定。【结果】在4 ℃时,mogR缺失后,菌株运动性显著强于亲本株(P<0.01),鞭毛合成量显著多于亲本株(P<0.001),鞭毛基因转录水平显著高于亲本株(P<0.001);缺失株ΔmogR的生长能力显著弱于亲本株(P<0.05)。回补株cΔmogR的运动能力、鞭毛合成量和鞭毛基因转录水平与亲本株相比,无显著性差异。【结论】在4 ℃时,Lm鞭毛产量少与MogR对鞭毛基因起转录抑制作用有关,低温条件下Lm能生长繁殖与MogR对鞭毛基因表达的抑制作用有关。本研究结果为揭示Lm低温生长机制提供了新的信息。     

拟态弧菌金属蛋白酶基因PrtV缺失株的构建及生物学特性分析

【背景】研究发现PrtV基因编码含多囊肾病(polycystic kidney disease)结构域的金属蛋白酶,其在多种细菌的致病过程中具有重要作用。拟态弧菌是一种感染多种水生动物的重要病原菌,PrtV基因在拟态弧菌致病中的作用尚不清楚。【目的】探究PrtV基因对拟态弧菌致病相关生物学特性的影响。【方法】采用自然转化的方法构建拟态弧菌PrtV基因缺失株(ΔPrtV),同时通过基因与质粒重组后电转化导入缺失株构建回补株(ΔPrtV/pPrtV),对突变株的生长特性、生化特征、生物被膜形成、自聚集能力、胞外产物卵磷脂酶和蛋白酶活性,以及致病性和细胞毒性等进行分析。【结果】与野生株相比,缺失株的生长特性、生物被膜形成、自聚集能力和卵磷脂酶活性无变化,但分解尿素、甘氨酸、香豆酸盐、鸟氨酸和赖氨酸的理化特性改变;胞外产物蛋白酶活性显著降低(P<0.05),细胞毒性显著下降(P<0.05),对杂交鲇的致病力下降10倍。【结论】PrtV基因与拟态弧菌的细胞毒性及致病性等多种生物学特性有关。该结果为进一步解析拟态弧菌PrtV基因功能及其致病机制提供了依据。     

单核细胞增多性李斯特菌谷胱甘肽合成酶调控鞭毛形成研究

[目的] 本试验旨在构建单核细胞增多性李斯特菌谷胱甘肽合成酶基因gshF缺失株和互补株并研究其在细菌运动和鞭毛形成中的调控作用。[方法] 采用同源重组的方法构建得到gshF缺失株后,测定野生株及缺失株的运动性和体外生长能力;同时利用荧光定量PCR方法检测gshF缺失株中鞭毛相关基因转录水平的变化。[结果] 缺失gshF后细菌在体外培养基中的生长能力未受影响,但缺失株的运动性及鞭毛形成能力显著降低。此外,缺失株中鞭毛形成重要调控基因gmaR以及鞭毛结构元件基因flaA的转录水平显著低于野生株,而其他鞭毛相关基因的转录水平变化不明显。[结论] 研究首次表明单增李斯特菌谷胱甘肽合成酶通过调控鞭毛重要基因的转录进而影响细菌的运动性和鞭毛形成,研究有助于深入理解重要食源性胞内菌通过精确调控鞭毛形成以适应外界和宿主环境的分子机制。     

单增李斯特菌毒力因子溶血素关键氨基酸位点在感染过程中的作用研究

【目的】探究单增李斯特菌溶血素O (listeriolysin O, LLO)中D3区域β8折叠片上第253位氨基酸(谷氨酰胺,Q)和第254位氨基酸(异亮氨酸,I)对单增李斯特菌(Listeria monocytogenes)感染生物学功能的影响。【方法】构建LLO_Q253A和LLO_I254A突变蛋白的原核表达菌株,以及利用同源重组方法构建hly_Q253A和hly_I254A突变株;通过表达纯化突变蛋白,测定溶血活性;比较LLO第253位Q和第254位I均突变成丙氨酸(A)后,对细菌体外生长能力、黏附侵袭、胞内迁移和增殖能力的影响。【结果】相应位点突变后,LLO蛋白均能够正常表达。在pH 6.5条件下,所有突变蛋白和突变株的溶血活性丧失。然而,在pH 5.5条件下,LLO_I254A和hly_I254A恢复了溶血活性。与野生株相比,突变株的体外生长、黏附能力和胞内增殖能力均无明显差异;突变株的侵袭能力和胞间迁移能力显著低于野生株。【结论】本研究证实第253位Q和第254位I均突变成A后,单增李斯特菌在pH 6.5条件下丧失溶血活性,并降低了感染宿主细胞的能力,但具体机制还有待进一步探索。本研究为深入探究LLO结构对单增李斯特菌生物学功能的影响奠定基础,对单增李斯特菌点突变株的构建具有一定参考意义。     

一株海洋来源铜绿假单胞菌Gxun-7角蛋白酶基因的克隆、表达及重组酶酶学性质

【背景】角蛋白酶是一类特异性降解角蛋白的水解酶,在动物饲料、生物肥料、医学、洗涤、制革及环境治理等方面具有重要的应用潜力。【目的】对前期从海洋环境筛选出的一株铜绿假单胞菌Gxun-7的角蛋白酶基因进行克隆、表达,并探究重组酶酶学性质,为角蛋白酶在工业生产中的应用奠定基础。【方法】以铜绿假单胞菌Gxun-7基因组推定的角蛋白酶基因为基础,设计引物克隆获得角蛋白酶基因kp2,构建重组表达质粒pET22b-kp2,并转化到E. coliRosettagamiB (DE3)中进行诱导表达,同时对重组表达菌株的表达条件进行优化。利用镍柱分离纯化重组角蛋白酶并研究其酶学性质。【结果】重组角蛋白酶的分子量约为33 kDa,最适温度和pH值分别为40 ℃和8.0,在温度30-60 ℃和pH 6.5-8.0具有较好的稳定性。金属离子Co²⁺、Cu²⁺和化学试剂十二烷基磺酸钠(sodium dodecyl sulfonate,SDS)、乙二胺四乙酸(ethylenediaminetetraacetic acid,EDTA)、苯甲基磺酰氟(phenylmethylsulfonyl fluoride,PMSF)对酶活力有抑制作用,而Mg²⁺、K⁺、巯基乙醇和二硫苏糖醇(dithiothreitol,DTT)对酶活力有促进作用。重组角蛋白酶具有良好的耐盐性,在12.5%的NaCl作用下相对酶活为87.55%。以酪蛋白为底物时,酶的K_m值为60.92 mg/mL、V_max值为9.70 U/mL。【结论】海洋来源铜绿假单胞菌Gxun-7的重组角蛋白酶具有良好的温度、碱、盐稳定性,可应用于工业生产中。     

617株耐亚胺培南铜绿假单胞菌耐药性及其产超广谱β-内酰胺酶基因型研究

探讨耐亚胺培南铜绿假单胞菌的耐药性及其产超广谱β-内酰胺酶基因型。收集2011年7月至2013年12月上海市中医药大学附属曙光医院临床分离的铜绿假单胞菌共1 125株,筛选亚胺培南耐药株,常规纸片法检测其耐药性,并用E-test检测金属β-内酰胺酶（MBL）,采用PCR法检测耐药基因型。结果显示,1 125株铜绿假单胞菌中耐亚胺培南铜绿假单胞菌共计617株,占54.8%;亚胺培南敏感铜绿假单胞菌共计508株,占45.2%。617株亚胺培南耐药铜绿假单胞菌100%为多重耐药,而亚胺培南敏感铜绿假单胞菌的多重耐药率仅为13.78%,明显较前者低（χ²=871.15,P<0.05）;亚胺培南耐药的铜绿假单胞菌中MBL表型阳性共126株,阳性率为15.4%,94株(74.60%)表现为VIM-2阳性,10株（7.94%）表现为IMP-1阳性,1株检出OXA-10,〖WTBZ〗且该例菌株同时表达VIM-2。临床分离的耐亚胺培南的铜绿假单胞菌多为多重耐药,其产MBL的主要基因型是VIM-2。     

铜绿假单胞菌lasI,rhlI基因功能缺陷株的构建和生物学功能验证

构建铜绿假单胞菌lasI,rhlI基因功能缺陷株,为进一步阐明氦氧饱和高气压暴露条件诱导lasI,rhlI基因介导铜绿假单胞菌毒力调节的分子机制研究奠定基础。用双亲株接合转移法删除lasI,rhlI基因ORF编码区,通过RT-PCR方法验证目标基因编码序列mRNA的缺失;通过对细菌生长增殖能力、弹性蛋白酶代谢活性和细菌绿脓菌素分泌能力等表型的测定,验证目标基因编码序列缺失后的基因调节功能的缺陷。结果表明成功构建铜绿假单胞菌lasI,rhlI基因功能缺陷株,可作为进一步研究的基因工程菌。     

Distribution of glucose/mannose-specific isolectins in pea (Pisum sativum L.) seedlings

We report on the distribution and initial characterization of glucose/mannose-specific isolectins of 4- and 7-d-old pea (Pisum sativum L.) seedlings grown with or without nitrate supply. Particular attention was payed to root lectin, which probably functions as a determinant of host-plant specificity during the infection of pea roots by Rhizobium leguminosarum bv. viciae. A pair of seedling cotyledons yielded 545±49 g of affinity-purified lectin, approx. 25% more lectin than did dry seeds. Shoots and roots of 4-d-old seedlings contained 100-fold less lectin than cotyledons, whereas only traces of lectin could be found in shoots and roots from 7-d-old seedlings. Polypeptides with a subunit structure similar to the precursor of the pea seed lectin could be demonstrated in cotyledons, shoots and roots. Chromatofocusing and isoelectric focusing showed that seed and non-seed isolectin differ in composition. An isolectin with an isoelectric point at pH 7.2 appeared to be a typical pea seed isolectin, whereas an isolectin focusing at pH 6.1 was the major non-seed lectin. The latter isolectin was also found in root cell-wall extracts, detached root hairs and root-surface washings. All non-seed isolectins were cross-reactive with rabbit antiserum raised against the seed isolectin with an isolectric point at pH 6.1. A protein similar to this acidic glucose/mannose-specific seed isolectin possibly represents the major lectin to be encountered by Rhizobium leguminosarum bv. viciae in the pea rhizosphere and at the root surface. Growth of pea seedlings in a nitrate-rich medium neither affected the distribution of isolectins nor their hemagglutination activity; however, the yield of affinity-purified root lectin was significantly reduced whereas shoot lectin yield slightly increased. Agglutination-inhibition tests demonstrated an overall similar sugar-binding specificity for pea seed and non-seed lectin. However root lectin from seedlings grown with or without nitrate supplement, and shoot lectin from nitrate-supplied seedlings showed a slightly different spectrum of sugar binding. The absorption spectra obtained by circular dichroism of seed and root lectin in the presence of a hapten also differed. These data indicate that nutritional conditions may affect the sugar-binding activity of non-seed isolectin, and that despite their similarities, seed and non-seed isolectins have different properties that may reflect tissue-specialization.Abbreviations IEF isoelectric focusing - MW molecular weight - pI isoelectric point - Psl1, Psl2 and Psl3 pea isolectins - SDSPAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis The authors wish to thank Professors L. Kanarek and M. van Poucke for helpful discussions.     

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     

Ectoenzymes in Prorocentrum minimum

Ectoenzymes, or enzymes associated with the cell-surface or periplasmic space, play an important role in organic matter cycling by rendering certain forms of dissolved organic matter bioavailable. Ectoenzyme activities may thereby help meet the nutritional demands of harmful algae such as Prorocentrum minimum. The activities of two ectoenzymes; leucine aminopeptidase and alkaline phosphatase, have been studied in axenic cultures of P. minimum. Leucine aminopeptidase releases non-polar amino acids such as leucine from the N-terminus of polypeptides, whereas alkaline phosphatase is an enzyme that is able to hydrolyze phosphate from phosphomonoesters. P. minimum alkaline phosphatase is the better studied of the two ectoenzymes and its characteristics are reviewed herein. Future research on P. minimum physiology will benefit from a growing suite of tools available for assessing the activity of alkaline phosphatase and other ectoenzymes in field populations and ultimately the work done with P. minimum will be useful for studies of other harmful species.     

PMA结合ddPCR检测食品中金黄色葡萄球菌的研究

研究了将叠氮溴化丙锭(PMA)与微滴式数字PCR(ddPCR)技术相结合,用于金黄色葡萄球菌活菌的检测。结果表明,强烈光照15 min,可以使PMA与死菌DNA共价交联,同时钝化游离的PMA;可以有效抑制金黄色葡萄球菌死菌DNAPCR扩增的PMA终浓度为2.0μg/m L;不抑制活菌DNA扩增的PMA最高浓度是5.0μg/m L。在不同死、活菌比例下,PMA-ddPCR可以定量检测活菌,避免了死菌DNA的干扰,本方法的检出限为10 copy/20μL。利用PMA-ddPCR检测人工污染鸡肉样品,最低可检出102cfu/m L的金黄色葡萄球菌。表明PMA-ddPCR方法的灵敏度高。     

Map position of Aldox (Aldehyde-oxidase) and Adh (Alcohol dehydrogenase) loci on autosome II of Musca domestica L.

The Aldox and Adh structural loci of Musca domestica L. belong to autosome II. They code for the enzymes aldehyde oxidase and alcohol dehydrogenase. Both these enzymes have allelic variants with specific electrophoretic mobility, which, on cellogel, are seen as single bands. The Aldox and Adh loci encompass a large map interval, which includes the morphological markers ar, cm, and car. The recombination frequencies between these five loci indicate the alignment Aldox-ar-cm-car-Adh.

---

Localisation chez Musca domestica L. des gènes Aldox et Adh codant les enzymes ald\;ehyde oxydase (AO) et alcool déshydrogénase (ADH)

Résumé Chez la mouche, les enzymes AO et ADH, observées par électrophorèse sur cellogel, présentent toutes 2 des formes alléliques exprimées par des bandes ayant une mobilité anodique propre.Les gènes structuraux Aldox et Adh, codant ces formes, sont liés entre eux et situés sur le chromosome 2. Ils se recombinent avec une fréquence élevée d'interchange; ils sont donc séparés par un intervalle important dans lequel sont compris les caractères visibles ar, cm, car. La fréquence des recombinaisons entre caractères visibles et gènes enzymatiques indique l'ordre suivant sur ce segment du deuxième chromosome de la mouche: Aldox, ar, cm, car, Adh.

     

3种外源诱导剂预处理对甜瓜抗病特征及其Fom 2和CHT基因表达的影响

该研究选用水杨酸(SA)、茉莉酸甲酯(MeJA)、Ca~(2+)、无菌水(对照)作为外源预处理诱导剂,以抗、感枯萎病甜瓜品种为材料,分别于诱导预处理2d后接种甜瓜枯萎菌,并于接种5、7、9d时观察发病情况,进行病情调查;在接种后1、3、5、7、9d取甜瓜叶片,分析抗病甜瓜(MR-1)和感病甜瓜(M1-15)叶片中甜瓜抗枯萎病基因(Fom-2)、几丁质酶基因(CHT)的表达变化,以探寻提高防治甜瓜枯萎病菌侵染的技术途径。结果显示:(1)外源MeJA和SA预处理接种后2品种的病情指数显著低于对照,但Ca~(2+)处理后的病情指数与对照无显著差异。(2)经外源诱导预处理接种后,MR-1和M1-15品种叶片的Fom-2和CHT基因均出现差异表达,但Ca~(2+)诱导其上调表达的效果微弱。(3)经SA、MeJA诱导预处理接种后,2品种叶片的Fom-2和CHT基因表达总体均显著高于对照;Fom-2基因的表达抗病甜瓜MR-1分别在接种后5d、7d时达到峰值,而感病甜瓜M1-15则均在接种9d时达到峰值;CHT基因的表达抗病甜瓜MR-1则均在接种后7d时达到峰值,而感病甜瓜M1-15分别在接种后7d、9d时达到峰值。(4)Ca~(2+)处理对抗、感甜瓜叶片的Fom-2和CHT基因的表达均无显著影响。(5)相关分析表明,经SA、MeJA诱导预处理接种后,甜瓜枯萎病病情指数与Fom-2和CHT基因表达量有显著的相关性;而Ca~(2+)处理效果不显著。研究表明:SA、MeJA通过诱导Fom-2、CHT基因上调表达,进而使甜瓜的抗病性提高,而Ca~(2+)处理对两基因表达和甜瓜抗病性均无显著影响。     

Differences between wheat and rice in the enzymic properties of ribulose-1,5-bisphosphate carboxylase/oxygenase and the relationship to photosynthetic gas exchange

The kinetic parameters of ribulose-1,5-bisphosphate (RuBP) carboxylase/oxygenase (EC 4.1.1.39) in wheat (Triticum aestivum L.) and rice (Oryza sativa L.) were determined by rapidly assaying the leaf extracts. The respective K _m and V _max values for carboxylase and oxygenase activities were significantly higher for wheat than for rice. In particular, the differences in the V _max values between the two species were greater. When the net activity of CO₂ exchange was calculated at the physiological CO₂-O₂ concentration from these kinetic parameters, it was 22% greater in wheat than in rice. This difference in the in-vitro RuBP-carboxylase/oxygenase activity between the two species reflected a difference in the CO₂-assimilation rate per unit of RuBP-carboxylase protein. However, there was no apparent difference in the CO₂-assimilation rate for a given leaf-nitrogen content between the two species. When the RuBP-carboxylase/oxygenase activity was estimated at the intercellular CO₂ pressure from the enzyme content and kinetic parameters, these estimated enzyme activities in wheat and rice were similar to each other for the same rate of CO₂ assimilation. These results indicate that the difference in the kinetic parameters of RuBP carboxylase between the two species was offset by the differences in RuBP-carboxylase content and conductance for a given leaf-nitrogen content.Abbreviations DTT dithiothreitol - EDTA ethylenediamine-tetraacetic - PAR photosynthetically active radiation - RuBP ribulose-1,5-bisphosphate     

The chlorophyll-a/b proteins of photosystem II in Chlamydomonas reinhardtii

We have adapted the procedure for the isolation of PSII membranes from higher plants (D.A. Berthold et al., 1981, FEBS Lett. 134, 231–234) to the green algae Chlamydomonas reinhardtii. The chlorophyll (Chl)-binding proteins from this PSII preparation have been further separated into single Chl-binding polypeptides and characterized spectroscopically. Seven single polypeptides were shown to bind Chl a and Chl b. In particular, we demonstrate that polypeptides p9, p10 and p22, which had not been previously shown to bind Chl a and b, have characteristics similar to those of CP29, CP26 and CP24 from higher plants. We note, however, that p9 and p10 are phosphorylatable in C. reinhardtii, at variance with CP29 and CP26 from higher plants. Our data support the notion that the PSII antenna systems in C. reinhardtii and in higher plants are very similar. Therefore, studies on the organization and regulation of light-harvesting processes in C. reinhardtii may provide information of general relevance for both green algae and higher plants.Abbreviations Chl chlorophyll - IEF isoelectrofocusing - LHC light harvesting complex - MW molecular weight - PAGE polyacrylamide gel electrophoresis - PS photosystem - RC reaction centre - SDS sodium dodecylsulfate We thank Dr. J. Olive (Institut Jacques Monod, Paris, France) for the electron-microscopy analysis, C. de Vitry (Institut de Biologie Physico-Chimique, Paris, France) for the kind gift of a PSII RC preparation and P. Dainese and M.L. Di Paolo (Universitá di Padova, Padova, Italy) for helpfull discussions. Professor Strasser and Elizbeth Scwartz (Université de Genova, Genova, Switzerland) are thanked for assistance in taking low-temperature fluorescence emission spectra. Roberto Bassi was recipient of a short-term fellowship from the European Molecular Biology Organization fellowship, during the early phases of the work.     

Production of l-lactic acid by electrodialysis fermentation (EDF)

An efficient process for producing l-lactic acid using an, EDF method is described. The results showed that intermittent EDF with continuous medium feed was the best one among the experiment methods employed. Comparing with the conventional EDF, intermittent EDF (seven on–off) with continuous medium feed indicated that the maximum value of o.d.₆₆₀ was not increased, but productivity was 1.5 times higher. The yield increased by above 30% and glucose transport decreased to 1/10 (from 0.46 to 0.05).