ABSTRACT
Bacterial esterases are gaining the importance in pharmaceuticals and agrochemical industries due to their excellent biocatalytic properties and a wide range of applications. In the present study, a novel gene encoding an esterase (designated as Est-CR) was identified from shotgun metagenomic sequencing data of camel rumen (Camelus dromedarius) liquor. The open reading frame consisted of 1,224bp, which showed 84.03% sequence identity to Bacteroidales bacterium, corresponding to a protein of 407 amino acids and has a catalytic domain belonging to an esterase. Est-CR belonged to family V with GLSMG domain. The purified enzyme with a molecular mass of 62.64 kDa was checked on SDS-PAGE, and its expression was confirmed by western blotting. The enzyme was active and stable over a broad range of temperature (35-65 °C), displayed the maximum activity at 50 °C and pH 7.0. Individually all metal ions inhibited the enzyme activity, while in combination, K2+, Ca2+, Mg2+ and Mn2+ metal ions enhanced the enzyme activity. The detergents strongly inhibited the activity, while EDTA (10 mM) increased the activity of the Est-CR enzyme. The enzyme showed specificity to short-chain substrates and displayed an optimum activity against butyrate ester. This novel enzyme might serve as a promising candidate to meet some harsh industrial processes enzymatic needs.
Subject(s)
Cations/chemistry , Esterases/chemistry , Metagenome/genetics , Metals/chemistry , Amino Acid Sequence , Animals , Bacteroides/genetics , Camelus , Catalytic Domain , Enzyme Activation , Enzyme Stability , Escherichia coli/genetics , Esterases/genetics , Esterases/metabolism , Hydrogen-Ion Concentration , Protein Binding , Rumen , Substrate Specificity , TemperatureABSTRACT
In addition to a wide variety of anaerobic and facultative anaerobic bacteria, camel rumen also harbors a diverse of eukaryotic organisms. In the present study, the eukaryotic communities of camel rumen were characterized using 18S rRNA amplicon sequencing. Metagenomic DNA was isolated from rumen samples of fourteen adult Bikaneri and Kachchhi breeds of camel fed different diets containing Jowar, Bajra, Maize, and Guar. Illumina sequencing generated 27,161,904 number of reads corresponding to 1543 total operational taxonomic units (OTUs). Taxonomic classification of community metagenome sequences from all the samples revealed the presence of 92 genera belonging to 16 different divisions, out of which Ciliophora (73%), Fungi (13%) and Streptophyta (9%) were found to be the most dominant. Notably, the abundance of Ciliophora was significantly higher in the case of Guar feed, while Fungi was significantly higher in the case of Maize feed, indicating the influence of cellulose and hemicellulose content of feedstuff on the composition of eukaryotes. The results suggest that the camel rumen eukaryotes are highly dynamic and depend on the type of diet given to the animal. Pearson's correlation analysis suggested the ciliate protozoa and fungi were negatively correlated with each other. To the best of our knowledge, this is first systematic study to characterize camel rumen eukaryotes, which has provided newer information regarding eukaryotic diversity patterns amongst camel fed on different diets.
