Uptake and metabolism of taurine in the green alga Chlorella fusca

Taurine entered the alga Chlorella fusca Shihira et Krauss strain 21l-8b via a pH and energy-dependent system ("permease"). Transport followed triphasic kinetics from 10⁻⁶ to 10⁻² M with K_m values for taurine of 5.4 × 10⁻⁵, 4.1 × l0⁻⁴ and l.5 × 10⁻³ M. This uptake system was specific for sulfonic acids and showed no affinity for α- and β -amino acids or Na⁺; thus the permease of C. fusca is different from all known taurine transport systems with respect to structural specificity and lack of Na⁺ -dependence. Uptake was not observed in sulfate-grown algae but developed as a response to sulfate limitation within 2 h. Sulfate addition caused a rapid decline in taurine transport capacity. Labeled taurine was rapidly metabolized in C. fusca to sulfate and ethanolamine, suggesting oxidative hydrolysis as the mechanism of C-S bond cleavage. Further incorporation of these catabolic products in C - and S -metabolism was demonstrated. Taurine catabolism was also detected in other green algae and some cyanobacteria.     

Changes in polyamines and related enzymes with loss of viability in rice seeds

Putrescine, spermidine and spermine of high vigour, low vigour and non-viable (classes 1, 2 and 3 respectively) seeds of Oryza sativa increased with loss of viability. The largest concentration of spermine was found in non-viable embryos. Spermine was absent in the husks of all the three categories of seeds. Arginine decarboxylase was greatest in high vigoured seeds and its activity gradually declined with loss of viability. However, diamine oxidase and polyamine oxidase activities gradually increased with the loss of viability of the seeds while DNA, RNA and protein contents decreased. The total content of polyamines increased on kinetin treatment but declined on ABA treatment. DNA, RNA and protein followed the same trend as polyamines. The polyamine contents increased by ca 3- and 4-fold, respectively, in high vigoured and low vigoured seeds on 10^?4 M kinetin treatment. The activity of ADC followed the same change as that of the polyamines in both cases, but the reverse was observed for the activities of diamine and polyamine oxidases.     

Arginine utilization by Chlorella autotrophica and Chlorella saccharophila

Chlorella saccharophila can utilize the amino acids arginine, glutamate. ornithine and proline as sole sources of nitrogen for growth. By comparison C. autotrophica utilized only arginine and ornithine. Following osmotic shock of Chlorella autotrophica from 50 to 150% artificial seawater rapid synthesis of proline (the main osmoregulatory solute in this alga) occurred in cells grown on arginine or citrulline. However, little proline synthesis occurred in ornithine-grown cells. Distribution of radiolabelled carbon from [¹⁴C]-arginine assimilation following osmotic shock of C. autotrophica agrees with the following pathway of arginine utilization: arginine→citrulline→ornithine→glutamate semialdehyde→pyrroline-5-carboxylate→proline. These 4 steps are catalysed by arginine deiminase (EC 3.5.3.6), citrullinase (EC 3.5.1.20), ornithine transaminase (EC 2.6.1.13) and pyrroline-5-carboxylate reductase (EC 1.5.1.2), respectively. Of these 4 enzymes, only arginine deiminase and pyrroline-5-carboxylate reductase were detected in the crude extract of the 2 Chlorella species. Arginine deiminase did not require specific cations for optimal activity. The deimi-nase showed maximal activity at pH 8.0 and followed Michaelis-Menten kinetics with an apparent K_m for L-arginine of 0.085 m M for the C. autotrophica enzyme and 0.097 m M for that of C. saccharophila. The activity of arginine deiminase was not influen-ced by growing C. saccharophila on arginine. Ornithine competitively inhibited arginine deiminase with an apparent K, of 2.4 m M for the C. autotrophica enzyme, and 3.8 m M for that of C. saccharophila . Arginine utilization by Chlorella is discussed in relation to that of other organisms.     

Antiatherogenic effect of a low lysine: arginine ratio of protein involves alteration in the aortic glycosamihoglycans and glycoproteins

The effect of alteration of lysine: arginine ratio of the protein on the aortic glycosaminoglycans and glycoproteins was studied in rats fed cholesterol free and atherogenic diet. The concentration of total glycosaminoglycans and of individual fractions was significantly lower in the aorta in the case of diet with lysine: arginine ratio of 1.0, than the diet with a ratio of 2.0. Rats fed globulin fraction isolated from sesame seeds, which has a lysine: arginine ratio of 0.67 also showed significantly lower concentration of total and individual glycosaminoglycan fractions in the aorta than those fed casein (lysine:arginine ratio 2.0). Concentration of total hexose and fucose in the glycoproteins was also lower in the aorta in the case of lysine: arginine ratio 1.0. These results in the light of previous reports of increase in the aortic glycosaminoglycans in the early stages of atherosclerosis and increase in the total hexose and fucose in the glycoproteins in the atherosclerotic aorta indicate that the antiatherogenic effect of a low lysine: arginine ratio in the protein involves alteration in the aortic glycosaminoglycans and glycoproteins.     

Low synonymous site variation at thelacY locus inEscherichia coli suggests the action of positive selection

We have determined the nucleotide sequences of sevenlacY alleles isolated from natural isolates ofEscherichia coli. Nucleotide heterozygosity estimates for this locus were compared to those obtained from previous studies of intraspecific variation at chromosomal loci, revealing thatlacY has unusually low synonymous site variation. The average pairwise heterozygosity of synonymous sites (Ks=0.0112+/-0.0100) is the second lowest reported and the lowest for loci that have an equivalent level of nonsynonymous variation. We consider several hypotheses to explain how different forces in evolution could act to create the observed pattern of polymorphism, including selection for translational efficiency and positive selection. Our analysis most strongly supports the hypothesis that positive selection has acted on thelacY locus inE. coli.     

Immunoreactivity between a monoclonal lupus autoantibody and the arginine/aspartic acid repeats within the U1-snRNP 70K autoantigen is conformationally restricted

Immunoreactivity of the arginine/aspartic acid (RD) repeats of the 70K protein of U1 small nuclear ribonucleoprotein (snRNP) was determined to be conformationally dependent. The monoclonal autoantibody 2.73, isolated from a lupus-prone MRL/n mouse model, is reactive with the RD repeat regions of U1 snRNP 70K protein. Immunochemical analysis of the antigenic determinants with use of chemically synthesized peptides characterized the 2.73 epitope as the RD repeat [Pelsue, S.,et al. (1993)Autoimmunity,15, 231–236] Analysis by circular dichroism (CD) and nuclear magnetic resonance spectroscopy indicates conformational preferences in the immunoreactive peptides. Computer analyses of CD spectra obtained on the RD-containing peptides predict-turns and-sheet to be the preferred conformations of the RD repeats. This structure was also predicted by the Chou-Fasman algorithm. The RD repeat is believed to be a conserved structural motif; however, the biological function is still unclear. Immunological and biochemical analysis of autoimmune antibodies and their respective antigenic determinants has helped to characterize the possible mechanisms that lead to autoimmune diseases. This is the first report of a conformationally dependent, linear epitope of an autoantibody.     

Sequence homology and structure predictions of the creatine kinase isoenzymes

Comparisons of the protein sequences and gene structures of the known creatine kinase isoenzymes and other guanidino kinases revealed high homology and were used to determine the evolutionary relationships of the various guamidino kinases. A CK framework is defined, consisting of the most conserved sequence blocks, and diagnostic boxes are identified which are characteristic for anyone creatine kinase isoenzyme (e.g. for vertebrate B-CK) and which may serve to distinguish this isoenzyme from all others (e.g. from M-CKs and Mi-CKs). Comparison of the guanidino kinases by near-UV and far-UV circular dichroism further indicates pronounced conservation of secondary structure as well as of aromatic amino acids that are involved in catalysis.Abbreviations GuaK guanidino kinase - CK creatine kinase - B-and M-CK brain and muscle cytosolic CK isoenzyme - Mi-CK mitochondrial CK isoenzyme - ArgK arginine kinase - Cr creatine - PCr phosphorylcreatine - PArg phosphorylarginine     

Polyamine metabolism during seedling development in rice

The main free amines identified during growth and development of rice seedlings were agmatine, putrescine, spermidine, diaminopropane and tyramine. Amine composition differed according to tissue and stages of development. Conjugated amines were only found in roots. We present evidence that arginine decarboxylase (ADC) regulates putrescine during the development of rice seedlings. When ADC action was blocked by DFMA (-DL-difluoromethylarginine, a specific irreversible inhibitor of ADC), polyamine titers and seedling development were diminished; when agmatine or putrescine was added, normal polyamine titers and growth were restored. The effects of DFMA were concentration dependent. DFMO (-DL-difluoromethylornithine, a specific irreversible inhibitor of ornithine decarboxylase or ODC) promoted growth and development at concentrations below 2 mM. This effect was probably related to its unexplained, but consistently observed slight enhancement of rice ADC. When the increase in the concentration of spermidine was prevented by CHA (cyclohexylammonium sulfate), the number of roots increased and the increase in length of leaves and roots was strongly inhibited. The addition of exogenous spermidine at the time of treatment with CHA reversed the inhibition by CHA.Abbreviations ADC arginine decarboxylase - ODC ornithine decarboxylase - DFMA -DL-difluoromethylarginine - DFMO -DL-difluoromethylornithine - CHA cyclohexylammonium sulfate     

Interactions of polyamines and nitrogen nutrition in plants

Biogenic amines occupy an important position among the many nitrogenous plant compounds. Polyamines are part of the overall metabolism of nitrogenous compounds, yet they do not seem to function in the 'normal' nitrogen nutrition. Rather, these widespread polycations (e. g. putrescine, spermidine and spermine) are involved in the regulation of growth and stress, probably by binding to negatively charged macromolecules. In addition, some diamines and polyamines are metabolized to yield 'secondary 'metabolites such as nicotine and other alkaloids. Previous studies have indicated that the ratio of nitrate to ammonium nutrition affects polyamine biosynthesis and content in intact plants. Thus, an increase in putrescine accumulation was found under conditions of excess ammonium ions, relative to nitrate. Modifications of nitrogen sources in the culture medium of tobacco cell suspensions (depletion of ammonium nitrate, or potassium nitrate, or both) resulted in marked changes in the content of cellular free polyamines. Considerable changes in the content of specific polyamines were also found with exposure to specific inhibitors of polyamine biosynthesis (difluoromethyl ornithine, difluoromethyl arginine, cyclohexylamine, methylglyoxal-bis-guanylhydrazone). However, a combination of nitrogen depletion of the medium and some inhibitors resulted in a very marked over-production of spermidine and spermine. The significance of these findings is discussed in relation to the assumption that polyamines act as a metabolic buffer, and maintain cellular pH under conditions where ammonium assimilation produces an excess of protons.     

Nitrate permease from Azotobacter chroococcum

Active transport systems in bacteria can be divided into two groups: those that are osmotic shock-resistant with one single membrane protein, and those that are shock-sensitive and have a membrane-bound protein complex plus a soluble periplasmic protein. Whether the bacterial assimilatory nitrate transport falls into the one or the other of these two groups has not been studied before. We report that nitrate uptake by the strictly aerobic, N₂-fixing heterotrophic bacterium Azotobacter chroococcum is sensitive to osmotic shock. The polypeptide composition of cytoplasmic membranes changes in response to the nitrogen source available to the cells. Incorporation of [³⁵S]-methionine into proteins as well as use of the A. chroococcum TRI mutant, which is defective in nitrate transport, and the A. choococcum MCD1 strain, a mutant unable to use nitrate as a nitrogen source, suggest that nitrate transport into A. chroococcum cells is mediated by a multicomponent system tightly bound to the cytoplasmic membrane.