Evidence for a close spatial location of the binding sites for CO2 and for photosystem II inhibitors

1. CO2-depletion of thylakoid membranes results in a decrease of binding affinity of the Photosystem II (PS II) inhibitor atrazine. The inhibitory efficiency of atrazine, expressed as I50-concentration (50% inhibition) of 2,6-dichlorophenolindophenol reduction, is the same in CO2-depleted as well as in control thylakoids. This shows that CO2-depletion results in a complete inactivation of a part of the total number of electron transport chains. 2. A major site of action of CO2, which had previously been located between the two electron acceptor quinone molecule B (or R) and Photosystem II inhibitor atrazine as suggested by the following observations: (a) CO2-depletion results in a shift of the binding constant (kappa b) of [14C]atrazine to thylakoid membranes indicating a decreased affinity of atrazine to membrane; (b) trypsin treatment, which is known to modify the Photosystem II complex at the level of B, strongly diminishes CO2 stimulation of electron transport reactions in CO2-depleted membranes; and (c) thylakoids from atrazine-resistant plants, which contain a Photosystem II complex modified at the inhibitor binding site, show an altered CO2-stimulation of electron flow. 3. CO2-depletion does not produce structural changes in enzyme complexes involved in Photosystem II function of thylakoid membranes, as shown by freeze-fracture studies using electron microscopy.     

Genetic studies among Ramgarhias and Ramdasias of Punjab, North India

This study reports results of investigations on ABO, Rh, haptoglobin and transferrin types in the two hitherto uninvestigated endogamous groups of Punjab, North India. Frequencies for the A, B, and O genes were found to be 0.1711, 0.2566, and 0.5723 in Ramdasias, and 0.1737, 0.2960, and 0.5303 in Ramgarhias. In all 13 Rhesus-negative individuals were encountered. The Hp2 allele shows a high frequency of 0.8060 for Ramdasias, and 0.7860 for Ramgarhias.     

A Nitrate Reductase-less Variant Isolated from Suspension Cultures of Datura innoxia (Mill.)

A comparative study has been carried out of the growth of two lines of Datura innoxia (Mill.) cells, designated DI-6 and NR1, their resistance to chlorate, and their ability to assimilate nitrate in sterile culture. The NR1 cell line was isolated from DI-6 cultures by first growing the latter in a nitrate-based medium for 5 days and then transferring the cells to a medium containing 2 grams liter⁻¹ of casein hydrolysate as the sole N source and 49 millimolar KClO₃ for a 6-week incubation period. Cells which survived the chlorate treatment then were transferred to casein hydrolysate medium and have been cultured in the absence of chlorate for more than 18 months (NR1).     

Changes in proline level in response to osmotic stress and exogenous amino acids in Raphanus sativus L. seedlings

The changes in endogenous proline levels of Raphanus sativus L. seedlings was monitored in presence of exogenous amino acids in normal and osmotically stressed seedlings. In unstressed seedlings, proline uptake was detected only at higher (1 mM) concentration of applied L-proline. however, proline uptake was promoted at all (1 μM to 1000 μM) concentrations of applied L-proline under osmotic stress conditions. Amongst other exogenous amino acids, L-leucine, L-glutamic acid, L-alanine, and L-histidine enhanced endogenous levels of proline, while exogenous hydorxyproline and γ-amino butyric acid reduced it.     

Experimental Absidia corymbifera infection in rabbits: Sequential pathological studies

Intravenous inoculation of young rabbits with 1.4 × 105 spores ml-1 of Absidia corymbifera produced gross and microscopic changes principally in the kidneys. Grossly, there was congestion, haemorrhages and greyish-white necrotic foci giving a mottled appearance to the kidneys. Microscopically, besides congestion and haemorrhages, there was a granulomatous reaction characterized by a central necrotic mass containing mycelium surrounded by a reactive zone of neutrophils, lymphocytes, a few macrophages and giant cells. Fibrous tissue proliferation was also present. Characteristic dichotomously branched non-septate fungal hyphae could be demonstrated in the sections of kidneys up to 14 days of infection. In one rabbit, which had died on the 8th day, small granulomatous lesions with fungal hyphae were noticed in the liver's parenchyma. This revised version was published online in June 2006 with corrections to the Cover Date.     

Temperature-induced production of recombinant human insulin in high-cell density cultures of recombinant Escherichia coli

The construction of expression vectors encoding either the human insulin A- or B-chains fused to a synthetic peptide and the temperature-induced expression of the recombinant genes in Escherichia coli are reported. Using this two-chain approach we also describe the separate isolation of the insulin A- and B-chains from inclusion bodies and their subsequent assembly into native human insulin. The production of the insulin fusion proteins were carried out in high-cell density fed-batch cultures using a synthetic medium with glucose as sole carbon and energy source. The expression of the recombinant genes by temperature-shift in high-cell density cultures of recombinant E. coli resulted in product yields of grams per litre of culture broth, e.g. 4.5 g of insulin B-chain fusion protein per litre of culture broth. This translates into an expression yield of about 800 mg of the insulin B-chain per litre of culture. Under similar cultivation conditions the expression yield of the insulin A-chain corresponds to approximately 600 mg per litre of culture. The metabolic burden imposed on the recombinant cells during temperature-induced production of insulin fusion proteins in high-cell density cultures is reflected in an increased respiratory activity and a reduction of the biomass yield coefficient with respect to glucose.     

Linking radiosilver to monoclonal antibodies reduced by ascorbic acia

Radiosilver-111 and Radiogold-199 were proposed by us (1) as suitable isotopes for radioimmunotherapy in areas such as India by reason of their suitable half-lives and B-emissions (Ag-111T _1/2=7.45 d and Au-199T _1/2=3.15 d). Since silver is monovalent, it is difficult to link to conventional bifunctional chelates. We therefore explored the use of sulfur-based linkers (2). Encouraged by the Thakur and De Fulvio Technique (3) of linking technetium to disulfide groups in antibodies reduced by ascorbic acid that is eminently biocompatible, we have explored the linkage of silver to immunoglobulin reduced by ascorbic acid. The linkage of silver was assessed with stable Ag-108 using dialysis to quantify the free silver after the reaction of silver and reduced immunoglobulins in various molar ratios (1∶1, 1∶2, 1∶5, 1∶10). The silver quantity was estimated gravimetrically after precipitation as chloride. It was observed that using these molar ratios there was negligible silver efflux into the dialysate, suggesting stable linkage. We also assessed the linkage using Ag-110M as radiotracer. The comparative results with the two techniques are described.     

Biomethanation of rice and wheat straw

When rice or wheat straw was added to cattle dung slurry and digested anaerobically, daily gas production increased from 176 to 331 l/kg total solids with 100% rice straw and to 194 l/kg total solids with 40% wheat straw. Not only was methane production enhanced by adding chopped crop residues but a greater biodegradability of organic matter in the straws was achieved.The authors are with the Microbiology and Molecular Genetics Unit, Tata Energy Research Institute, 158 Jor Bagh, New Delhi 110 003, India     

Purification and characterization of annexin proteins from bovine lung

Calcium-dependent association with a detergent-extracted particulate fraction was used as the first step in the purification of a group of phospholipid binding proteins. Elution of the detergent-insoluble fraction with excess ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) resulted in the release of several soluble proteins, termed calcium-activated proteins or CAPs. In the present paper, we describe the simultaneous purification of these CAPs and characterize their interaction with phospholipid, actin, and calmodulin. Partial sequence analysis has identified the majority of the CAPs as members of the annexin family of calcium and phospholipid binding proteins. Two additional CAPs may be novel proteins, one of which appears to be an annexin protein. All CAPs demonstrated Ca2(+)-dependent binding to phosphatidylserine vesicles but did not bind to phosphatidylcholine vesicles. The majority of CAPs exhibited Ca2(+)-dependent binding to F-actin; however, only CAP-III affected the rate of conversion of G-actin to F-actin. The interaction of CAP-III and lipocortin-85 with F-actin resulted in a Ca2(+)-dependent increase in both light scattering and sedimentation of F-actin under comparatively low centrifugal force. In contrast, only lipocortin-85 caused the formation of F-actin bundles. Although all of the CAPs bound to a calmodulin affinity column in a Ca2(+)-dependent manner, attempts to demonstrate binding of CAPs to native calmodulin were unsuccessful. These studies therefore document the similar behavior of the CAPs toward phospholipid and calmodulin but clearly show that F-actin binding or bundling is not a general property of these proteins. The reported purification procedure should allow further comparative studies of these proteins.