Functional characterization and target discovery of glycoside hydrolases from the digestome of the lower termite Coptotermes gestroi

Background

Lignocellulosic materials have been moved towards the forefront of the biofuel industry as a sustainable resource. However, saccharification and the production of bioproducts derived from plant cell wall biomass are complex and lengthy processes. The understanding of termite gut biology and feeding strategies may improve the current state of biomass conversion technology and bioproduct production.

Results

The study herein shows comprehensive functional characterization of crude body extracts from Coptotermes gestroi along with global proteomic analysis of the termite's digestome, targeting the identification of glycoside hydrolases and accessory proteins responsible for plant biomass conversion. The crude protein extract from C. gestroi was enzymatically efficient over a broad pH range on a series of natural polysaccharides, formed by glucose-, xylose-, mannan- and/or arabinose-containing polymers, linked by various types of glycosidic bonds, as well as ramification types. Our proteomic approach successfully identified a large number of relevant polypeptides in the C. gestroi digestome. A total of 55 different proteins were identified and classified into 29 CAZy families. Based on the total number of peptides identified, the majority of components found in the C. gestroi digestome were cellulose-degrading enzymes. Xylanolytic enzymes, mannan- hydrolytic enzymes, pectinases and starch-degrading and debranching enzymes were also identified. Our strategy enabled validation of liquid chromatography with tandem mass spectrometry recognized proteins, by enzymatic functional assays and by following the degradation products of specific 8-amino-1,3,6-pyrenetrisulfonic acid labeled oligosaccharides through capillary zone electrophoresis.

Conclusions

Here we describe the first global study on the enzymatic repertoire involved in plant polysaccharide degradation by the lower termite C. gestroi. The biochemical characterization of whole body termite extracts evidenced their ability to cleave all types of glycosidic bonds present in plant polysaccharides. The comprehensive proteomic analysis, revealed a complete collection of hydrolytic enzymes including cellulases (GH1, GH3, GH5, GH7, GH9 and CBM 6), hemicellulases (GH2, GH10, GH11, GH16, GH43 and CBM 27) and pectinases (GH28 and GH29).     

Seed germination temperatures of eight Mexican Agave species with economic importance

The genetic diversity of Agave plants is threatened by clonal commercial reproduction and climatic change. Sexual reproduction is uncommon and research on seed germination is scarce. The present study evaluated the seed germination of Agave lechuguilla, Agave striata, Agave americana var. marginata, Agave asperrima, Agave cupreata, Agave duranguesis, Agave angustifolia ssp. tequilana and Agave salmiana at constant temperatures (10, 15, 20, 25, 30, 35 and 40°C). Initial imbibition (after the first 12 h) was significantly variable among species, positively correlated with seed weight (r = 0.6560, P < 0.001) and increased with temperature (from 35% at 10°C to 66% at 40°C). Temperature affected maximum imbibition (83–150%) for A. asperrima, A. lechuguilla, A. salmiana and A. striata; other species averaged 110%. Most germination kinetics best fitted a logistic model, whereas only a few treatments fit a Weibull model. The time to germination onset diminished (P < 0.05) from 125–173 h at 15°C to 68–84 h at 25°C, and then ascended to 84–196 h at 35°C. The mean germination rate and seed germination percentage after 312 h peaked at 25°C (0.50–0.95% seeds/h and 85–99%, respectively) and fell (P < 0.05) to near zero at 10 and 40°C. Temperatures of 10, 35 and 40°C were partially lethal to A. asperrima, A. duranguensis and A. salmiana seeds. The time to germination onset, seed germination percentage after 312 h and mean germination rate are best described by a Gaussian distribution, with its optimum at approximately 25°C. Thus, optimum temperatures are related to the ecological characteristics of each species area.     

Conversion of Woodward's Reagent K in an aqueous medium: mathematical analysis applied to enzyme modification

A change in the optical density of Woodward's Reagent K solution at 340 nm has been shown. It is observed after the reagent has been dissolved in a weakly acidic medium. The optical density correlates with the reagent's ability to inhibit transketolase. A method for assay of the inhibitor concentration changes in the medium during enzyme modification is suggested.     

Tobacco Transformants Bearing Antisense Suppressor of Proline Dehydrogenase Gene, Are Characterized by Higher Proline Content and Cytoplasm Osmotic Pressure

The antisense suppressor was constructed for proline dehydrogenase gene (PDH; a fragment of PDH from Arabidopsis in antisense orientation and under the control of 35S promoter of cauliflower mosaic virus, CMV). In Nicotiana tabacum SR1 tobacco transformants bearing antisense suppressor for PDH, the proline content and the cytoplasm osmotic pressure were increased. The proline content in these transformants varied, whereas cytoplasm osmotic pressure was stable, which seems to reflect complicated relationships between these characteristics of the plant cell.     

Chorioallantoic placentation in Galea spixii (Rodentia,Caviomorpha, Caviidae)

Background  

Placentas of guinea pig-related rodents are appropriate animal models for human placentation because of their striking similarities to those of humans. To optimize the pool of potential models in this context, it is essential to identify the occurrence of characters in close relatives.     

Electron temperature in nitrogen afterglow: Dependence of theoretical results on the adopted set of cross sections and on the type of molecular distribution over vibrational levels

The electron energy distribution function in nitrogen afterglow is simulated using different available sets of cross sections for electron scattering by vibrationally excited molecules. The calculations are performed for two types of molecular distribution over vibrational levels, namely, for the Boltzmann and Treanor-Gordiets distributions. It is shown that the calculated values of the electron temperature in nitrogen afterglow depend strongly on the set of cross sections used and on the type of molecular distribution over vibrational levels. The validity of comparison between theoretical and experimental results is discussed.     

Alternative translation start sites and hidden coding potential of eukaryotic mRNAs

It is widely suggested that a eukaryotic mRNA typically contains one translation start site and encodes a single functional protein product. However, according to current points of view on translation initiation mechanisms, eukaryotic ribosomes can recognize several alternative translation start sites and the number of experimentally verified examples of alternative translation is growing rapidly. Also, the frequent occurrence of alternative translation events and their functional significance are supported by the results of computational evaluations. The functional role of alternative translation and its contribution to eukaryotic proteome complexity are discussed.     

Computer System mRNA-FAST (mRNA Function,Activity, STructure)

Computer system mRNA-FAST (mRNA Function, Activity, STructure; http://wwwmgs.bionet.nsc.ru/mgs/dbases/trsig/) is described. The system has been developed to analyze nucleotide sequences of mRNA and to measure their essential properties. The system compiles the data base on translation signals including nucleotide sequences of the regulatory regions with structural and experimental information on their specific activities. It also contains programs to search for local homology between mRNA and translation signals, to search for potential signals basing on analysis of the oligonucleotide dictionaries, and to model secondary RNA structure. Possible applications of the system mRNA-FAST are discussed.     

ABC: software for interactive browsing of genomic multiple sequence alignment data

Background  

Alignment and comparison of related genome sequences is a powerful method to identify regions likely to contain functional elements. Such analyses are data intensive, requiring the inclusion of genomic multiple sequence alignments, sequence annotations, and scores describing regional attributes of columns in the alignment. Visualization and browsing of results can be difficult, and there are currently limited software options for performing this task.