Changes in leucyl-tRNAs and aminoacyl tRNA synthetases in developing and aging soybean cotyledons

Leucine specific tRNA of soybean cotyledons was frationated into six peaks (1–6). The relative amounts of Leu-tRNA 5 and 6 are lower in developing cotyledons than in germinating cotyledons. Leu-tRNA synthetase from developing cotyledons is less active in aminoacylating Leu-tRNA 5 and 6 compared to enzyme from 5-day-old germinating cotyledons. Leu-tRNA synthetase from cotyledons of germinating seedlings and developing cotyledons can be fractionated into three peaks (1–3). Peak 1 in the developing cotyledon is about 36% less than peak 1 from 5-day-old germinating cotyledons. Peaks 2 and 3 from developing cotyledons are about 10 and 18% higher than from germinating cotyledons, respectively. Peak 1 from developing cotyledons acylates all six species of Leu-tRNA in contrast with peak 1 from germinating cotyledons, which essentially acylates only Leu-tRNA 5 and 6. The specificity of peaks 2 and 3 towards Leu-tRNA 1–4 is identical in both the organs.     

In vitro removal of human IgG autoantibodies by affinity filtration using immobilized L-histidine onto PEVA hollow fiber membranes

Histidine was immobilized onto PEVA membrane to obtain an affinity support for human IgG removal from serum with a view to clinical apheresis for the treatment of autoimmune diseases. These membranes were able to remove in vitro several autoantibodies from the serum of SLE patients.     

Natural and chemically induced oligomeric ribonucleases: structural study by immobilized metal ion affinity electrophoresis and their functional relationship

Oligomerization can endow proteins with novel structural and catalytic properties. The native dimer of bovine seminal ribonucleases (BS-RNase) binds, melts and catalyses the hydrolysis of double-stranded ribonucleic acids 30-fold better than its pancreatic homologue, the monomeric RNase A. Chemically induced oligomers of pancreatic RNase A are also found to show an increase in enzyme activity on double-stranded poly(A).poly(U) (Libonati, M. Bertoldi, M. and Sorrentino, S. (1996) Biochem. J. 318, 287-290) and, therefore, can be considered as potential immunosuppressive and cytotoxic agents. We report here a study on the relationship between surface histidine topography in oligomeric forms of these ribonucleases and their catalytic properties. Subtle changes in structure conformation of both BS-RNase and oligomeric RNase A are shown to result in a modification of the affinity of these proteins toward the immobilized transition-metal chelate, IDA-Cu(II). Because, such conformational change has been shown to correlate with an improvement of the newly acquired biological activities upon oligomerization, we can conclude that surface histidines topography constitutes an exquisite probe for the study of protein structure/function relationship.     

Carbohydrate metabolic pathway genes associated with quantitative trait loci (QTL) for obesity and type 2 diabetes: Identification by data mining

Increasing consumption of refined carbohydrates is now being recognized as a primary contributor to the development of nutritionally related chronic diseases such as obesity and type 2 diabetes mellitus (T2DM). A data mining approach was used to evaluate the role of carbohydrate metabolic pathway genes in the development of obesity and T2DM. Data from public databases were used to map the position of the carbohydrate metabolic pathway genes to known quantitative trait loci (QTL) for obesity and T2DM and for examining the pathway genes for the presence of sequence and structural genetic variants such as single nucleotide polymorphisms (SNPs) and copy number variants (CNS), respectively. The results demonstrated that a majority of the genes of the carbohydrate metabolic pathways are associated with QTL for obesity and many for T2DM. In addition, some key genes of the pathways also encode non-synonymous SNPs that exhibit significant differences in population frequencies. This study emphasizes the significance of the metabolic pathways genes in the development of disease phenotypes, its differential occurrence across populations and between individuals, and a strategy for interpreting an individuals' risk for disease.     

Circadian rhythms in myocardial metabolism and contractile function: influence of workload and oleate

Multiple extracardiac stimuli, such as workload and circulating nutrients (e.g., fatty acids), known to influence myocardial metabolism and contractile function exhibit marked circadian rhythms. The aim of the present study was to investigate whether the rat heart exhibits circadian rhythms in its responsiveness to changes in workload and/or fatty acid (oleate) availability. Thus, hearts were isolated from male Wistar rats (housed during a 12:12-h light-dark cycle: lights on at 9 AM) at 9 AM, 3 PM, 9 PM, and 3 AM and perfused in the working mode ex vivo with 5 mM glucose plus either 0.4 or 0.8 mM oleate. Following 20-min perfusion at normal workload (i.e., 100 cm H(2)O afterload), hearts were challenged with increased workload (140 cm H(2)O afterload plus 1 microM epinephrine). In the presence of 0.4 mM oleate, myocardial metabolism exhibited a marked circadian rhythm, with decreased rates of glucose oxidation, increased rates of lactate release, decreased glycogenolysis capacity, and increased channeling of oleate into nonoxidative pathways during the light phase. Rat hearts also exhibited a modest circadian rhythm in responsiveness to the workload challenge when perfused in the presence of 0.4 mM oleate, with increased myocardial oxygen consumption at the dark-to-light phase transition. However, rat hearts perfused in the presence of 0.8 mM oleate exhibited a markedly blunted contractile function response to the workload challenge during the light phase. In conclusion, these studies expose marked circadian rhythmicities in myocardial oxidative and nonoxidative metabolism as well as responsiveness of the rat heart to changes in workload and fatty acid availability.     

Optimization, purification and characterization of novel thermostable, haloalkaline, solvent stable protease from Bacillus halodurans CAS6 using marine shellfish wastes: a potential additive for detergent and antioxidant synthesis

A protease producing marine bacterium, Bacillus halodurans CAS6 isolated from marine sediments, was found to produce higher enzyme by utilizing shrimp shell powder. Optimum culture conditions for protease production were 50 °C, pH 9.0, 30 % NaCl and 1 % shrimp shell powder (SSP) and the protease purified with a specific activity of 509.84 U/mg. The enzyme retained 100 % of its original activity even at 70 °C, pH 10.0 and 30 % NaCl for 1 h. The purified protease exhibited higher stability when treated with ionic, non-ionic (72–94 %) and commercial detergents (76–88 %), and organic solvents (88–126 %). Significant blood stain removal activity was found with the enzyme in washing experiments. The culture supernatant supplemented with 1 % SSP showed 93.67 ± 2.52 % scavenging activity and FT-IR analysis of the reaction mixture confirmed the presence of antioxidants such as cyclohexane and cyclic depsipeptide with aliphatic amino groups. These remarkable qualities found with this enzyme produced by Bacillus halodurans CAS6 could make this as an ideal candidate to develop the industrial process for bioconversion of marine wastes and antioxidant synthesis.     

Histidine mapping of serine protease: a synergic study by IMAC and molecular modelling

The immobilized metal ion affinity (IMA) interaction of different serine proteases, namely porcine and bovine trypsins and BPN' and Carlsberg subtilisins, was studied on Sepharose-IDA-CuII. Both trypsins were resolved into their different subspecies, whereas the subtilisins appeared as only one species. The use of diethyl pyrocarbonate-modified enzymes demonstrated the contribution of histidine(s) as the sole interacting site(s). The use of different peptidic and chemical inhibitors complexed to the enzymes confirmed the contribution of histidine(s) as the interacting site(s) and further resulted in different chromatographic patterns for the free and complexed serine proteases. Comparison of the chromatographic data for each enzyme with the accessible surface area calculation by molecular modelling on the available crystallographic structure allowed us to hypothesize a map of the surface-accessible histidine on each enzyme.