BackgroundSerratiopeptidase is an alkaline metalloendopeptidase, which acquired wide significance because of its therapeutic applications. The present study was undertaken for purification, characterization, and structural elucidation of serratiopeptidase produced from Streptomyces hydrogenans var. MGS13.ResultThe crude enzyme was purified by precipitating with ammonium sulfate, dialysis, and Sephadex gel filtration, resulting in 34% recovery with a 12% purification fold. The purified enzyme S.AMP13 was spotted as a single clear hydrolytic band on casein zymogram and whose molecular weight was found to be 32 kDa by SDS-PAGE. The inhibitor and stability studies revealed that this enzyme is metalloprotease, thermostable, and alkaline in nature. The maximum serratiopeptidase activity was observed at 37 °C and pH 9.0. The partial amino acid sequence of the purified enzyme S.AMP13 by LC-MS/MS analysis shows the closest sequence similarities with previously reported alkaline metalloendopeptidases. The amino acid sequence alignment of S.AMP13 shared a conserved C-terminus region with peptidase-M10 serralysin superfamily at amino acid positions 128–147, i.e., ANLSTRATDTVYGFNSTAGR revealed that this enzyme is a serralysin-like protease. The kinetic studies of the purified enzyme revealed a Km of 1 mg/mL for its substrate casein and Vmax of 319 U/mL/min. The 3D structure of the purified enzyme was modeled by using SWISS-MODEL, and the quality of the structure was authenticated by assessing the Ramachandran plot using PROCHECK server, which suggested that the enzyme was stable with good quality.ConclusionInhibitor, stability, electrophoretic, and bioinformatic studies suggested that the purified enzyme obtained from S. hydrogenans var. MGS13 is a serralysin-like protease.Electronic supplementary materialThe online version of this article (10.1186/s43141-019-0002-7) contains supplementary material, which is available to authorized users.
Nageswara et al.: Statistical Optimization and Biological Characterization of Serralysin Serralysin is well known to exhibit anti-inflammatory and fibrinolytic properties. The current research designed a cost-effective serralysin production medium from Streptomyces hydrogenans var. MGS13 with the aid of solid state fermentation. Four pre-screened factors, namely horse gram flour concentration, inoculum size, initial moisture content and soya bean meal were modeled by central composite design for optimizing in order to predict their influence on serralysin production. Analysis of variance results showed a high coefficient of determination (R 2 ) value of 0.9611, ensuring a satisfactory adjustment of the quadratic model with the experimental data and F value 26.45 (p value of ˂0.0001) indicated that the model was significant. The design of experiment assisted production process enhanced 1.3 fold productivity at the best possible conditions consisting 5.0 g of horse gram flour, 1.2 ml of inoculum (1×10 6 CFU/ml), 44 % of initial moisture content and soya bean meal 1.0 % w/w. Besides this study, the in vitro fibrinolytic and antiinflammatory activities were carried with purified serralysin of Streptomyces hydrogenans var. MGS13. The results revealed that the purified enzyme exhibited fibrinolytic and anti-inflammatory activity in a dose dependent manner. Further one-way analysis of variance and Dunnett's multiple comparisons statistically justify the data p<0.05 in both activities.
Objective: In the present study, we have discussed the antibacterial activity of methanolic extract of Grewia tiliaefolia Vahl leaf and the isolation of pure phytochemicals by using column chromatography.
Methods: The powdered material was extracted with methanol using a soxhlet extractor and isolation of compounds by column chromatography. Evaluation of antibacterial activity determined the zone of inhibition by disc diffusion method.
Results: Three compounds were isolated from methanolic extract of the leaf of Grewia tiliaefolia using column chromatography, namely daucosterol, lupeol, and friedelin were purified after isolation and characterized by using UV, FTIR, NMR, MASS spectroscopic techniques, and antimicrobial activity was carried out on the crude methanolic extract a higher zone of inhibition was observed against Staphylococcus aureus with a zone diameter of 34.1±0.513 mm at a concentration of 10 mg/ml.
Conclusion: The results observed in this research work conclude that the methanolic extract showed good antibacterial activity may be due to the compounds isolated like daucosterol, lupeol, and friedelin.
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