ABSTRACT
The evolution, survival, and adaptation of microbes are consequences of gene duplication, acquisition, and divergence in response to environmental challenges. In this context, enzymes play a central role in the evolution of organisms, because they are fundamental in cell metabolism. Here, we analyzed the enzymatic repertoire in 6,467 microbial genomes, including their abundances, and their associations with metabolic maps. We found that the enzymes follow a power-law distribution, in relation to the genome sizes. Therefore, we evaluated the total proportion enzymatic classes in relation to the genomes, identifying a descending-order proportion: transferases (EC:2.-), hydrolases (EC:3.-), oxidoreductases (EC:1.-), ligases (EC:6.-), lyases (EC:4.-), isomerases (EC:5.-), and translocases (EC:7-.). In addition, we identified a preferential use of enzymatic classes in metabolism pathways for xenobiotics, cofactors and vitamins, carbohydrates, amino acids, glycans, and energy. Therefore, this analysis provides clues about the functional constraints associated with the enzymatic repertoire of functions in Bacteria and Archaea.
ABSTRACT
The luteinizing hormone receptor (LHR) is a glycoprotein member of the G protein-coupled receptors superfamily. It participates in corpus luteum formation and ovulation in females and acts in testosterone synthesis and spermatogenesis in males. In this study, we extracted RNA from sheep testicles and synthetized the cDNA to amplify the gene lhr-bed. This gene consists of 762 bp and encodes 273 amino acids of the extracellular domain of LHR. The lhr-bed was cloned into pJET1.2/blunt, then subcloned into pCOLD II, and finally, transformed in E. coli BL21 (DE3) cells. Because the induced rLHR-Bed protein was found in the insoluble fraction, we followed a modified purification protocol involving induction at 25 °C, subjection to denaturing conditions, and on-column refolding to increase solubility. We confirmed rLHR-Bed expression by means of Western blot and mass spectrometry analysis. It is currently known that the structure stem-loop 5'UTR on pCOLD II vector is stable at 15 °C. We predicted and obtained RNAfold stability at 25 °C. We successfully obtained the recombinant LHR extracellular domain, with protein yields of 0.2 mg/L, and purity levels of approximately 90%, by means of a single chromatographic purification step. The method described here may be used to obtain large quantities of rLHR-Bed in the future.
ABSTRACT
Phage therapy consists of applying bacteriophages, whose natural function is to kill specific bacteria. Bacteriophages are safe, evolve together with their host, and are environmentally friendly. At present, the indiscriminate use of antibiotics and salt minerals (Zn2+ or Cu2+) has caused the emergence of resistant strains that infect crops, causing difficulties and loss of food production. Phage therapy is an alternative that has shown positive results and can improve the treatments available for agriculture. However, the success of phage therapy depends on finding effective bacteriophages. This review focused on describing the potential, up to now, of applying phage therapy as an alternative treatment against bacterial diseases, with sustainable improvement in food production. We described the current isolation techniques, characterization, detection, and selection of lytic phages, highlighting the importance of complementary studies using genome analysis of the phage and its host. Finally, among these studies, we concentrated on the most relevant bacteriophages used for biocontrol of Pseudomonas spp., Xanthomonas spp., Pectobacterium spp., Ralstonia spp., Burkholderia spp., Dickeya spp., Clavibacter michiganensis, and Agrobacterium tumefaciens as agents that cause damage to crops, and affect food production around the world.