Purification and characterization of moderately thermostable raw-starch digesting α-amylase from endophytic Streptomyces mobaraensis DB13 associated with Costus speciosus.

Endophytic actinobacteria are known to produce various enzymes with potential industrial applications. Alpha-amylase is an important class of industrial enzyme with a multi-dimensional utility. The present experiment was designed to characterize a moderately thermostable α-amylase producing endophytic Streptomyces mobaraensis DB13 isolated from Costus speciosus (J. Koenig) Sm. The enzyme was purified using 60% ammonium sulphate precipitation, dialysis, and Sephadex G-100 column chromatography. Based on 12% SDS-PAGE, the molecular weight of the purified α-amylase was estimated to be 55 kDa. The maximum α-amylase activity was achieved at pH 7.0, 50°C and it retained 80% of its activity at both pH 7.0 and 8.0 after incubation for 2 h. The α-mylase activity is strongly enhanced by Ca2+, Mg2+, and inhibited by Ba2+. The activity remains stable in the presence of Tween-80, SDS, PMSF, and Triton X-100; however, ß-mercaptoethanol, EDTA, and H2O2 reduced the activity. The kinetic parameters Km and Vmax values for this α-amylase were calculated as 2.53 mM and 29.42 U/mL respectively. The α-amylase had the ability to digest various raw starches at a concentration of 10 mg/mL at pH 7.0, 50°C, where maize and rice are the preferred substrates. The digestion starts after 4 h of incubation, which reaches maximum after 48 h of incubation. These results suggest that S. mobaraensis DB13 is a potential source of moderately thermostable α-amylase enzyme, that effciently hydrolyzes raw starch. It suggesting that this α-amylase is a promising candidate to be use for industrial purposes.

Characterization and purification of esterase from Cellulomonas fimi DB19 isolated from Zanthoxylum armatum with its possible role in diesel biodegradation.

Endophytic bacteria inhabit all or part of their life cycle within the tissues of healthy plants, without causing any apparent symptoms of disease. They are treasure trove of several hydrolytic enzymes with distinct characteristics. Esterase is one of such enzymes and this study aims to characterize esterase produced by endophytic actinobacteria Cellulomonas fimi DB19 isolated from Zanthoxylum armatum with its capacity to degrade diesel oil. The enzyme was purified with purification fold 8.22 and specific activity 124.72 U/mg with 16.43% recovery. The purified enzyme showed a single protein band on SDS-PAGE having molecular mass of approximately 39 kDa. The Km and Vmax value for p-nitrophenyl acetate were 2.23 mM and 22.04 U/mL, respectively. The enzyme was stable in the pH range 6-9 with its optimal activity at pH 8.0. The enzyme was stable at 40 °C and retained more than 80% activity after incubation for two h. The enzyme activity was positively influenced in the presence of Na+, Ba2+, Ca2+, and negatively by Mn2+, and Mg2+. The EDTA and PMSF inhibited the enzyme activity and retained its activity in the presence of SDS, H2O2, ß-mercaptoethanol, and organic solvents. Application of the isolate in degradation of diesel showed that its growth and degradation capacity enhanced in media supplemented with 0.2-4% of diesel oil with maximum at 3% of diesel oil. Furthermore, esterase activity was greater in media containing diesel than control which is suggesting the plausible role of esterase produced by Cellulomonas fimi DB19 in the degradation of diesel oil.

Seasonal Variation Influence Endophytic Actinobacterial Communities of Medicinal Plants from Tropical Deciduous Forest of Meghalaya and Characterization of Their Plant Growth-Promoting Potentials.

The endophytic actinobacteria constitute a diverse community which has vast potential importance that may be exploited in pharmaceutical, agricultural, and biotechnological industries. However, the effects of seasonal changes on distribution of endophytic actinobacteria in medicinal plants of Meghalaya are largely uncharacterized. Here, we investigated host and seasonal influence on diversity of endophytic actinobacteria residing in roots of six medicinal plant species of Meghalaya. A total of 493 cultivable endophytic actinobacterial isolates representing 41 species were obtained from root segments of six plant species which had been collected during four different seasons of 2011-2012 and 2012-2013. Among the host plant species, maximum actinobacterial colonization was observed in Costus speciosus and minimum in Potentilla fulgens. In regard to seasons, the highest actinobacterial colonization and relative abundance were observed during summer season and least was recorded during the winter season. It was ascertained that though endophytic actinobacteria have varying capacity to colonize in different plant species during the seasons, colonization is not found to be species-specific. Culture-independent attempt also inferred that actinobacterial community varied amongst the six medicinal plants during the different seasons. Hence, seasons are influential factors in the colonization capacity of endophytic actinobacterial community. Furthermore, plant growth-promoting activities were recorded in 34.15% of the isolates. Hence, these results indicate that endophytic actinobacteria from the selected medicinal plants also represent an important source of plant growth-promoting bioactive metabolites.