A primordial tRNA modification required for the evolution of life?

The evolution of reading frame maintenance must have been an early event, and presumably preceded the emergence of the three domains Archaea, Bacteria and Eukarya. Features evolved early in reading frame maintenance may still exist in present-day organisms. We show that one such feature may be the modified nucleoside 1-methylguanosine (m(1)G37), which prevents frameshifting and is present adjacent to and 3' of the anticodon (position 37) in the same subset of tRNAs from all organisms, including that with the smallest sequenced genome (Mycoplasma genitalium), and organelles. We have identified the genes encoding the enzyme tRNA(m(1)G37)methyltransferase from all three domains. We also show that they are orthologues, and suggest that they originated from a primordial gene. Lack of m(1)G37 severely impairs the growth of a bacterium and a eukaryote to a similar degree. Yeast tRNA(m(1)G37)methyltransferase also synthesizes 1-methylinosine and participates in the formation of the Y-base (yW). Our results suggest that m(1)G37 existed in tRNA before the divergence of the three domains, and that a tRNA(m(1)G37)methyltrans ferase is part of the minimal set of gene products required for life.     

Marfey’s reagent for chiral amino acid analysis: A review

Summary. The present paper describes characteristics and application of Marfeys reagent (MR) including general protocols for synthesis of the reagent and diastereomers along with advantages, disadvantages and the required precautions. Applications, and comparison with other derivatizing agents, for the resolution of complex mixtures of DL-amino acids, amines and non-proteinogenic amino acids, peptides/amino acids from microorganisms, cysteine residues in peptides, and evaluation of racemizing characteristics have been discussed. Separation mechanisms of resolution of amino acid diastereomers and replacement of Ala–NH₂ by suitable chiral moieties providing structural analogs and different chiral variants and their application as a derivatizing agent to examine the efficiency, and reactivity of the reagent have been focussed. Use of MR for preparing CSPs for direct enantiomeric resolution has also been included.On leave from Department of Chemistry, Indian Institute of Technology Roorkee, Roorkee 247 667, India.     

Is alcoholic acidic silver nitrate reagent really specific for the histochemical localization of ascorbic acid?

Summary The histochemical localization of ascorbic acid in plant tissues with the alcoholic acidic silver nitrate reagent is shown here to be not specific for ascorbic acid, since some of the polyphenolic substances, including flavonoids, which are known to be widely distributed in plant tissues, are also able to reduce the acidic alcoholic silver nitrate reagent at low temperature (0–4°C) and at pH 2 to 2.5 in dark. This method may perhaps be used for animal tissues where flavonoid pigments do not occur in such large quantities as they do in plants. I therefore, come to the inevitable conclusion that the use of alcoholic acidic silver nitrate reagent in localizing ascorbic acid in plant tissues may be highly misleading.     

A role for PPARα in the regulation of arginine metabolism and nitric oxide synthesis

The pleiotropic effects of PPARα may include the regulation of amino acid metabolism. Nitric oxide (NO) is a key player in vascular homeostasis. NO synthesis may be jeopardized by a differential channeling of arginine toward urea (via arginase) versus NO (via NO synthase, NOS). This was studied in wild-type (WT) and PPARα-null (KO) mice fed diets containing either saturated fatty acids (COCO diet) or 18:3 n-3 (LIN diet). Metabolic markers of arginine metabolism were assayed in urine and plasma. mRNA levels of arginases and NOS were determined in liver. Whole-body NO synthesis and the conversion of systemic arginine into urea were assessed by using ¹⁵N₂-guanido-arginine and measuring urinary ¹⁵NO₃ and [¹⁵N]-urea. PPARα deficiency resulted in a markedly lower whole-body NO synthesis, whereas the conversion of systemic arginine into urea remained unaffected. PPARα deficiency also increased plasma arginine and decreased citrulline concentration in plasma. These changes could not be ascribed to a direct effect on hepatic target genes, since NOS mRNA levels were unaffected, and arginase mRNA levels decreased in KO mice. Despite the low level in the diet, the nature of the fatty acids modulated some effects of PPARα deficiency, including plasma arginine and urea, which increased more in KO mice fed the LIN diet than in those fed the COCO diet. In conclusion, PPARα is largely involved in normal whole-body NO synthesis. This warrants further study on the potential of PPARα activation to maintain NO synthesis in the initiation of the metabolic syndrome.     

Modelling the reaction course of N-acetylneuraminic acid synthesis from N-acetyl-d-glucosamine—new strategies for the optimisation of neuraminic acid synthesis

In this work, a model describing the complete enzyme catalysed synthesis of N-acetylneuraminic acid (Neu5Ac) from N-acetyl-d-glucosamine (GlcNAc) is presented. It includes the combined reaction steps of epimerisation from GlcNAc to N-acetyl-d-mannosamine (ManNAc) and the aldol condensation of ManNAc with sodium pyruvate yielding Neu5Ac. The model is expedient to predict the reaction course for various initial and feed concentrations and therefore to calculate reaction times and yields. The equilibrium constants calculated from the kinetic constants via the Haldane relationship correspond with experimental values very well (0.26 calculated and 0.24 experimental value for the epimerisation, 27.4 l mol⁻¹ calculated and 28.7 l mol⁻¹ experimental for the aldol condensation). The actual relevance of the model is shown by a scale-up. Using the model, an optimisation of reaction conditions in consideration of different targets is possible. Exemplarily, it is presented how the optimal ratio of the two enzymes in the reaction can be determined and how the composition of the reaction solution in a fed-batch reactor can be designed to meet downstream processing needs.     

A novel insight into the cost–benefit model for the evolution of botanical carnivory

Background The cost–benefit model for the evolution of botanical carnivory provides a conceptual framework for interpreting a wide range of comparative and experimental studies on carnivorous plants. This model assumes that the modified leaves called traps represent a significant cost for the plant, and this cost is outweighed by the benefits from increased nutrient uptake from prey, in terms of enhancing the rate of photosynthesis per unit leaf mass or area (A_N) in the microsites inhabited by carnivorous plants.Scope This review summarizes results from the classical interpretation of the cost–benefit model for evolution of botanical carnivory and highlights the costs and benefits of active trapping mechanisms, including water pumping, electrical signalling and accumulation of jasmonates. Novel alternative sequestration strategies (utilization of leaf litter and faeces) in carnivorous plants are also discussed in the context of the cost–benefit model.Conclusions Traps of carnivorous plants have lower A_N than leaves, and the leaves have higher A_N after feeding. Prey digestion, water pumping and electrical signalling represent a significant carbon cost (as an increased rate of respiration, R_D) for carnivorous plants. On the other hand, jasmonate accumulation during the digestive period and reprogramming of gene expression from growth and photosynthesis to prey digestion optimizes enzyme production in comparison with constitutive secretion. This inducibility may have evolved as a cost-saving strategy beneficial for carnivorous plants. The similarities between plant defence mechanisms and botanical carnivory are highlighted.     

A novel modeling approach for the “end-to-end” analysis of marine ecosystems

There is a growing demand for “end-to-end” models, which are modeling tools used to analyze and understand the fundamental complexities of marine ecosystems and processes emerging from the interaction of individuals from different trophic groups with respect to the physical environment and, even, human activity. These models are valuable quantitative tools for ecosystem-based management. To explore potential answers to complex questions regarding ecosystems using these models, it is necessary to incorporate classical ontogenic changes through the life cycle of target individuals, in addition to inherited behavioral strategies, as an additional differentiating aspect, particularly when the behavior has a direct impact on the ecosystem phenomena under study. However, it is difficult to combine different fine scale time and spatial granularities to infer animal behavior and ontogenic development. This complexity has kept these two levels of analysis separated, because most current tools do not have the required computational resources and advanced software architecture. To address this issue, we propose an individual-based modeling framework that is capable of handling and unifying the two experimental categories with a comprehensive biological and behavioral model that strictly adheres to the physiological functions of ingestion, growth, and metabolism of organisms. In addition, this model incorporates the exchange and transfer of mass and energy through local interactions at all trophic levels (lower to higher), the physical environment, and anthropogenic activity. For the framework to model short time events, such as classical predator–prey interactions, while also generating long-term ecosystem emergent properties, a special interleaving scheduling engine and physical space computer model was devised, which optimizes memory and processing resources. The framework was tested through several experiments with a three-population ecosystem containing up to 40 thousand organisms evolving inside a 200,000 m² simulation environment during 12,000 model-hours; yet, requiring only a few hours of program execution on a regular personal computer. The model included various environmental physical elements, such as several hundred shelters, the number of which can be easily modified in each experiment to simulate substrate degradation and its impact on populations. With the aid of the quantitative and qualitative tools provided by the model, it was possible to observe a coupling between prey and predator population dynamics. In conclusion, we confirmed that the end-to-end model developed here could successfully generate detailed specific hypotheses about fish behavior and quantify impacts on population dynamics.     

Crystal structure of shrimp arginine kinase in binary complex with arginine—a molecular view of the phosphagen precursor binding to the enzyme

Arginine kinase (AK) is a key enzyme for energetic balance in invertebrates. Although AK is a well-studied system that provides fast energy to invertebrates using the phosphagen phospho-arginine, the structural details on the AK-arginine binary complex interaction remain unclear. Herein, we determined two crystal structures of the Pacific whiteleg shrimp (Litopenaeus vannamei) arginine kinase, one in binary complex with arginine (LvAK-Arg) and a ternary transition state analog complex (TSAC). We found that the arginine guanidinium group makes ionic contacts with Glu225, Cys271 and a network of ordered water molecules. On the zwitterionic side of the amino acid, the backbone amide nitrogens of Gly64 and Val65 coordinate the arginine carboxylate. Glu314, one of proposed acid–base catalytic residues, did not interact with arginine in the binary complex. This residue is located in the flexible loop 310–320 that covers the active site and only stabilizes in the LvAK-TSAC. This is the first binary complex crystal structure of a guanidine kinase in complex with the guanidine substrate and could give insights into the nature of the early steps of phosphagen biosynthesis.     

Network genomics – A novel approach for the analysis of biological systems in the post-genomic era

Network Genomics studies genomics and proteomics foundations of cellular networks in biological systems. It complements systems biology in providing information on elements, their interaction and their functional interplay in cellular networks. The relationship between genomic and proteomic high-throughput technologies and computational methods are described, as well as several examples of specific network genomic application are presented.     

Survey of phytopathogenic pseudomonads for a restriction and modification system active on the double-stranded ribonucleic acid phage ϕ6

A total of 380 pseudomonad strains from 39 nomenspecies and 41 strains from 7 other bacterial genera were screened for a double-stranded ribonucleic acid modification and restriction system using the double-stranded ribonucleic acid bacteriophage 6. Of these 421 strains, 8 showed the low plating efficiency (10^–5 to 10^–7) characteristic of such a system. Howver, the phage propagated in 7 of the 8 were host-range mutants; the remaining strain showed some characteristics of a host-modification system but the results were equivocal.     

Synthesis of chlorophyll–amino acid conjugates as models for modification of proteins with chromo/fluorophores

A chlorophyll-a derivative bonded directly with epoxide at the peripheral position of the chlorin π-system was reacted with N-urethane and C-ester protected amino acids bearing an alcoholic or phenolic hydroxy group as well as a carboxy group at the residue to give chlorophyll–amino acid conjugates. The carboxy residues of N,C-protected aspartic and glutamic acids were esterified with the epoxide in high yields. The synthetic conjugates in dichloromethane had absorption bands throughout the visible region including intense red-side Qy and blue-side Soret bands. By their excitation at the visible bands, strong and efficient fluorescence emission was observed up to the near-infrared region. The chromo/fluorophores are promising for preparation of functional peptides and modification of proteins.     

Design and development of novel p-aminobenzoic acid derivatives as potential cholinesterase inhibitors for the treatment of Alzheimer’s disease

Based on the quantitative structure-activity relationship (QSAR), some novel p-aminobenzoic acid derivatives as promising cholinesterase enzyme inhibitors were designed, synthesized, characterized and evaluated to enhance learning and memory. The in vitro enzyme kinetic study of the synthesized compounds revealed the type of inhibition on the respective acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes. The in vivo studies of the synthesized compounds exhibited significant reversal of cognitive deficits in the animal models of amnesia as compared to standard drug donepezil. Further, the ex vivo studies in the specific brain regions like the hippocampus, hypothalamus, and prefrontal cortex regions also exhibited AChE inhibition comparable to standard donepezil. The in silico molecular docking and dynamics simulations studies of the most potent compound 22 revealed the consensual interactions at the active site pocket of the AChE.     

Chemical modification of lipase with ferromagnetic modifier — A ferromagnetic-modified lipase

Summary A ferromagnetic modifier was prepared by reacting ferrous(Fe²⁺)- and ferric(Fe³⁺)-ions with polyethylene glycol having two carboxyl groups (MW:2000) at pH 8.0–8.5. Lipase fromPseudomonas fragi 22–39B was coupled with the modifier using water-soluble carbodiimide. The modified lipase, which was dispersed into buffered solutions in the size range of 30–70 nm, exerted the hydrolytic activity of 8.0 U/mg. In a magnetic field of 250 Oe, the ferromagnetic-modified lipase was readily recovered from the colloidal solution.     

A novel glucuronoyl esterase from Aspergillus fumigatus—the role of conserved Lys residue in the preference for 4-O-methyl glucuronoyl esters

Applied Microbiology and Biotechnology - Cellulose in plant cell walls is mainly covered by hemicellulose and lignin, and thus efficient removal of these components is thought to be a key step in...     

Methylation of calmodulin at carboxylic acid residues in erythrocytes. A non-regulatory covalent modification?

The physiological role of protein carboxy-group methylation reactions in human erythrocytes was studied with calmodulin as an endogenous methyl-group acceptor. The steady-state degree of calmodulin carboxy-group methylation is substoichiometric both in intact cells and in a lysed-cell system (about 0.0003 mol of methyl groups/mol of polypeptide). Purified erythrocyte calmodulin is a substrate for a partially purified erythrocyte carboxy-group methyltransferase and can be methylated to the extent of about 0.0007-0.001 mol of methyl groups/mol of polypeptide. This erythrocyte protein methyltransferase displays an apparent specificity for atypical racemized and/or isomerized D-aspartate and L-isoaspartate residues [McFadden & Clarke (1982) Proc. Natl. Acad. Sci. U.S.A. 79, 2460-2464; Murray & Clarke (1984) J. Biol. Chem. 259, 10722-10732]. Exposure of calmodulin to elevated temperatures before methylation results in racemization of aspartate and/or asparagine residues, and may result in isoaspartate formation as well. The methylatability of these samples also increases as a function of time of heating, independent of the pH (over the range pH 5-9) or Ca2+ concentration; the most significant increase occurs during the initial 60 min, when calmodulin retains a fraction of its biological activity. These results are consistent with the hypothesis that methylation of calmodulin may occur at these uncommon aspartate residues, but are not consistent with a regulatory role for the methylation reaction.