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     

Structure of a stabilizing disulfide bridge mutant that closes the active-site cleft of T4 lysozyme.

The engineered disulfide bridge between residues 21 and 142 of phage T4 lysozyme spans the active-site cleft and can be used as a switch to control the activity of the enzyme (Matsumura, M. & Matthews, B.W., 1989, Science 243, 792-794). In the oxidized form the disulfide increases the melting temperature of the protein by 11 degrees C at pH 2. The crystal structure of this mutant lysozyme has been determined in both the reduced and oxidized forms. In the reduced form, the crystal structure of the mutant is shown to be extremely similar to that of wild type. In the oxidized form, however, the formation of the disulfide bridge causes the alpha-carbons of Cys 21 and Cys 142, on opposite sides of the active-site cleft, to move toward each other by 2.5 A. In association with this movement, the amino-terminal domain of the protein undergoes a rigid-body rotation of 5.1 degrees relative to the carboxy-terminal domain. This rotation occurs about an axis passing through the junction of the amino-terminal and carboxy-terminal domains and is also close to the axis that best fits the apparent thermal motion of the amino-terminal domain seen previously in crystals of wild-type lysozyme. Even though the engineered Cys 21-Cys 142 disulfide links together the amino-terminal and carboxy-terminal domains of T4 lysozyme, it does not reduce the apparent mobility of the one domain relative to the other. The pronounced "hinge-bending" mobility of the amino-terminal domain that is suggested by the crystallographic thermal parameters of wild-type lysozyme persists in the oxidized (and reduced) mutant structures. In the immediate vicinity of the introduced disulfide bridge the mutant structure is more mobile (or disordered) than wild type, so much so that the exact conformation of Cys 21 remains obscure. As with the previously described disulfide bridge between residues 9 and 164 of T4 lysozyme (Pjura, P.E., Matsumura, M., Wozniak, J.A., & Matthews, B.W., 1990, Biochemistry 29, 2592-2598), the engineered cross-link substantially enhances the stability of the protein without making the folded structure more rigid.     

Daily and seasonal variation in water relations of macchia shrubs and trees in France (Montpellier) and Turkey (Antalya)

Based upon two different research studies in the mediterranean regions of France and Turkey, drought resistance strategies were investigated in a broad group of species. The diurnal and seasonal patterns of the water relations of different lifeforms from the thermo-mediterranean to submediterranean lifezones were compared. Three sites near Montpellier, in Southern France, and five sites near Antalya, Turkey were used for this comparison. Xylem pressure potential and relative stomatal aperture were the key water relations parameters collected in France while these parameters as well as osmotic potential and leaf conductance were studied in Turkey.From the 26 different study species investigated in France, 7 distinct types of stomatal control were observed, with the deciduous lifeforms showing the least control, the sclerophyllous and coniferous evergreens the greatest control and the malacophyllous shrublets intermediate levels of control. Predawn water potential values provided a means of classifying species according to their temporal and spatial utilization of site water reserves. The comparison of turgor potentials (difference between water and osmotic potentials) gave an insight into leaf adaptations to site moisture. Species with high predawn water potentials generally maintain positive turgor even at midday during the summer, whereas species with low predawn values were frequently at zero turgor even at predawn. Phlomis grandiflora was the most extreme species with mid-summer predawns and midday water potentials of –6 MPa and osmotic potentials never more negative than –2.4 MPa.     

phlb基因诱导小麦ABD染色体组部分同源染色体配对的研究

通过花药培养首次获得了“中国春”phlb突变体单倍体,同时也获得了“中国春”单倍体。对其细胞学观察表明,前者花粉母细胞减数分裂中期Ⅰ每个细胞染色体交叉为5.08个,后者为1.30个。证明了phlb基因在单倍体状态下具有强的诱导ABD染色体组部分同源染色体间的配对作用。     

Production of wheat-barley recombinant chromosomes through induced homoeologous pairing

Summary Sears' phlb mutant was used successfully for the first time to induce pairing and recombination between specific barley chromosomes and their wheat homoeologues. Pairing was induced in specially constructed genetic stocks having 19 pairs of wheat chromosomes and triply monosomic for either barley chromosome arm 6HL or 3HL, a related wheat chromosome, and chromosome 5B of wheat carrying the phlb mutation. Wheat-barley recombinant chromosomes were isolated from among the progeny obtained from self-fertilization of the triple monosomic stocks, by screening for dissociation of biochemical markers on the barley arms. Glutamic oxaloacetic transaminase (GOT), aconitase hydratase (ACO), and dipeptidase (DIP) isozymes were used to select recombinants involving the 6HL arm, and esterase (EST) and malate dehydrogenase (MDH) were used for the 3HL arm. Altogether, six recombinants involving 6HL (1.4%) and six involving 3HL (1.1%) were isolated. These wheat-barley recombinant chromosomes are being used to construct a detailed gene order map of barley based on biochemical and molecular markers.     

几种担子菌胞外漆酶同工酶谱的研究

从猴头(Hericium erinaceus)、亚侧耳(Hohenbuehelia serotina)、毛柄金钱菌(Co-llybia velutipes)、多脂鳞伞(Pholiota adiposa)、香菇(Lentinus edodes)、黑木耳(Auriculariaauricula)、草菇(Volvaria volpacea)二孢蘑菇(Agaricus bisporus)和紫孢侧耳(Pleurotussapidus)、侧耳(Pleurotus ostreatus)风尾菇(P.sajor-caju)、金顶侧耳(P.citrinopileatus)、鲍鱼菇(P.cystidiosus)、佛罗里达侧耳(P.florida)、爪哇漏斗状侧耳(P.sajor-caju var.javanicus)商品化菌种的培养滤液里所得到的胞外漆酶(LC),采用聚丙烯酰胺凝胶等电聚焦盘状电泳方法,分析比较了它们的胞外漆酶同工酶。经多批次比较试验,获得了侧耳属七个种和八个不同属间各自恒定的漆酶同工酶谱。在这些酶谱中,分别包含了 A 组(pH3—5)和 B 组(pH5—8)的谱带。而且,所有菌种间漆酶带的等电点值(pI)各有异同。没有测定菌种间各漆酶带含量差别。讨论了漆酶同工酶谱用于分类研究的可能性。