BACKGROUND： Nattokinase （NK） is a serine protease enzyme of the subtilisin family. It exhibits a strong fibrinolytic activity. The fibrinolytic enzymes from Bacillus sp. have attracted interest as thrombolytic agen...BACKGROUND： Nattokinase （NK） is a serine protease enzyme of the subtilisin family. It exhibits a strong fibrinolytic activity. The fibrinolytic enzymes from Bacillus sp. have attracted interest as thrombolytic agents because of their efficiency in the fibrinolytic process including plasmin activation. METHODS： In the present study, VIT garden soil was collected and subjected to isolation process in order to screen for the NK production. Screening for NK enzyme was performed by radial caseinolytic assay. The production of NK enzyme was done in two different production medium for comparative studies. The NK enzyme was purified by gel permeation chromatography. The activity of the purified NK was checked by clot lysis and casein digestion assay. To investigate the structural basis of NK and fibrinogen interaction and also to identify the best binding mode, molecular dynamics and docking studies were performed. RESULTS： Based on the morphological and biochemical characterization, the isolate was identified as Bacillus sp. The overall purification fold of NK was about 3 with the specific activity of 664U/mg and 9.9% yield. Homogeneity of the purified enzyme was analyzed and confirmed by the single band obtained in SDS-PAGE. Molecular weight of the purified protease was estimated as 25 kDa. Purified NK enzyme exhibited 97% of effective clot lysis activity. The NK was docked in to the knob region of the fibrinogen at its binding site using Dock server. A total of 26 residues of fibrinogen and 29 residues of NK constitute the interface region. However, 9 residues offibrinogen （THR238, MET264, LYS266, ARG275, THR277, ALA279, ASN308, MET310, and LYS321） and 8 residues ofNK （GLY61, SER63, THR99, PHE189, LEU209, TYR217, ASN218, and MET222） are involved in intact binding. CONCLUSIONS： A significant amount of NK enzyme was obtained from Bacillus sp. The docking analysis revealed that the NK and fibrinogen adopt an extended binding pattern and interacts with the crucial residues to exhibit their activity.展开更多
BACKGROUND： Serratia is one of the most important groups of bacteria which produces proteolytic enzymes effectively and known to possess anti-inflammatory properties. The main focus of the current study was to extrac...BACKGROUND： Serratia is one of the most important groups of bacteria which produces proteolytic enzymes effectively and known to possess anti-inflammatory properties. The main focus of the current study was to extract the enzyme serratiopeptidase and pigment prodigiosin from Serratia mascescens. Prodigiosin is a red colored pigment produced by the bacterium Serratia marcescens. It is emerging as a valuable molecule because of its large applications. It has already been proved that pigmented strain of Serratia marcescens is less virulent than non-pigmented strains. Moreover the strain we have obtained is from farm soil which indicates that prodigiosin production can be carded safely using this strain. METHODS： In the present study, the isolate VITASP strain was confirmed by morphological, biochemical and molecular studies. The enzyme and pigment were analyzed for anti-oxidant, anti-inflammatory and cytotoxic properties. RESULTS： The isolate was further confirmed and identified as Serratia marcescens with 99% similarity. The extracted pigment showed potent radical scavenging effect with 86% and the enzyme was found to inhibit 83%, which was significant in comparison to ascorbic acid standard. The in vitro anti-inflammatory effect of pigment in controlled experimental conditions revealed its protection at 88% and the enzyme with 90%. Aspirin was used as the reference drug. The present findings exhibited a concentration dependent inhibition. The cytotoxic bioassay of pigment showed the IC5o value as （50） ~tg/mL with 63% cytotoxicity which was statistically significant compared to positive control. CONCLUSIONS： Therefore, it appears to be an essential remedial and application research. It may turn out to be highly beneficial to mankind in solving many problems associated with human health.展开更多
BACKGROUND： Marine actinomycetes are efficient producers of new secondary metabolites that show different biological activities, including antibacterial, antifungal, anticancer, insecticidal, and enzyme inhibition ac...BACKGROUND： Marine actinomycetes are efficient producers of new secondary metabolites that show different biological activities, including antibacterial, antifungal, anticancer, insecticidal, and enzyme inhibition activities. METHODS： The morphological, physiological, and biochemical properties of the strain Streptomyees sp. VITPSA were confirmed by conventional methods. Antibacterial, anti-oxidant, anti-inflammatory, anti-diabetic, and cytotoxic activities of Streptomyces sp. VITPSA extract were determined. The media were optimized for the production of secondary metabolites. Characterization and identification of secondary metabolites were conducted by high-performance liquid chromatography, gas chromatography-mass spectroscopy, and Fourier transform infrared spectroscopy analysis. RESULTS： The strain showed significant antibacterial, anti-oxidant, and cytotoxic activities, moderate anti-inflammatory activity, and no satisfactory anti-diabetic activity. The ethyl acetate extract of Streptomyces sp. VITPSA showed maximum antibacterial activity against two gram-positive and gram-negative bacteria at 0.5 mg/mL. The antioxidant potential of the crude extract exhibited strong reducing power activity at 0.5 mg/mL with 95.1% inhibition. The cytotoxic effect was found to be an ICso of 50 gg/mL on MCF-7 cell lines. Experimental design of optimization by one-factor analysis revealed the most favorable sucrose, yeast extract, pH （7.25）, and temperature （28~C） conditions for the effective production of secondary metabolites. CONCLUSION： This study revealed that Streptomyces sp. VITPSA is an excellent source of secondary metabolites with various bioactivities.展开更多
文摘BACKGROUND： Nattokinase （NK） is a serine protease enzyme of the subtilisin family. It exhibits a strong fibrinolytic activity. The fibrinolytic enzymes from Bacillus sp. have attracted interest as thrombolytic agents because of their efficiency in the fibrinolytic process including plasmin activation. METHODS： In the present study, VIT garden soil was collected and subjected to isolation process in order to screen for the NK production. Screening for NK enzyme was performed by radial caseinolytic assay. The production of NK enzyme was done in two different production medium for comparative studies. The NK enzyme was purified by gel permeation chromatography. The activity of the purified NK was checked by clot lysis and casein digestion assay. To investigate the structural basis of NK and fibrinogen interaction and also to identify the best binding mode, molecular dynamics and docking studies were performed. RESULTS： Based on the morphological and biochemical characterization, the isolate was identified as Bacillus sp. The overall purification fold of NK was about 3 with the specific activity of 664U/mg and 9.9% yield. Homogeneity of the purified enzyme was analyzed and confirmed by the single band obtained in SDS-PAGE. Molecular weight of the purified protease was estimated as 25 kDa. Purified NK enzyme exhibited 97% of effective clot lysis activity. The NK was docked in to the knob region of the fibrinogen at its binding site using Dock server. A total of 26 residues of fibrinogen and 29 residues of NK constitute the interface region. However, 9 residues offibrinogen （THR238, MET264, LYS266, ARG275, THR277, ALA279, ASN308, MET310, and LYS321） and 8 residues ofNK （GLY61, SER63, THR99, PHE189, LEU209, TYR217, ASN218, and MET222） are involved in intact binding. CONCLUSIONS： A significant amount of NK enzyme was obtained from Bacillus sp. The docking analysis revealed that the NK and fibrinogen adopt an extended binding pattern and interacts with the crucial residues to exhibit their activity.
文摘BACKGROUND： Serratia is one of the most important groups of bacteria which produces proteolytic enzymes effectively and known to possess anti-inflammatory properties. The main focus of the current study was to extract the enzyme serratiopeptidase and pigment prodigiosin from Serratia mascescens. Prodigiosin is a red colored pigment produced by the bacterium Serratia marcescens. It is emerging as a valuable molecule because of its large applications. It has already been proved that pigmented strain of Serratia marcescens is less virulent than non-pigmented strains. Moreover the strain we have obtained is from farm soil which indicates that prodigiosin production can be carded safely using this strain. METHODS： In the present study, the isolate VITASP strain was confirmed by morphological, biochemical and molecular studies. The enzyme and pigment were analyzed for anti-oxidant, anti-inflammatory and cytotoxic properties. RESULTS： The isolate was further confirmed and identified as Serratia marcescens with 99% similarity. The extracted pigment showed potent radical scavenging effect with 86% and the enzyme was found to inhibit 83%, which was significant in comparison to ascorbic acid standard. The in vitro anti-inflammatory effect of pigment in controlled experimental conditions revealed its protection at 88% and the enzyme with 90%. Aspirin was used as the reference drug. The present findings exhibited a concentration dependent inhibition. The cytotoxic bioassay of pigment showed the IC5o value as （50） ~tg/mL with 63% cytotoxicity which was statistically significant compared to positive control. CONCLUSIONS： Therefore, it appears to be an essential remedial and application research. It may turn out to be highly beneficial to mankind in solving many problems associated with human health.
文摘BACKGROUND： Marine actinomycetes are efficient producers of new secondary metabolites that show different biological activities, including antibacterial, antifungal, anticancer, insecticidal, and enzyme inhibition activities. METHODS： The morphological, physiological, and biochemical properties of the strain Streptomyees sp. VITPSA were confirmed by conventional methods. Antibacterial, anti-oxidant, anti-inflammatory, anti-diabetic, and cytotoxic activities of Streptomyces sp. VITPSA extract were determined. The media were optimized for the production of secondary metabolites. Characterization and identification of secondary metabolites were conducted by high-performance liquid chromatography, gas chromatography-mass spectroscopy, and Fourier transform infrared spectroscopy analysis. RESULTS： The strain showed significant antibacterial, anti-oxidant, and cytotoxic activities, moderate anti-inflammatory activity, and no satisfactory anti-diabetic activity. The ethyl acetate extract of Streptomyces sp. VITPSA showed maximum antibacterial activity against two gram-positive and gram-negative bacteria at 0.5 mg/mL. The antioxidant potential of the crude extract exhibited strong reducing power activity at 0.5 mg/mL with 95.1% inhibition. The cytotoxic effect was found to be an ICso of 50 gg/mL on MCF-7 cell lines. Experimental design of optimization by one-factor analysis revealed the most favorable sucrose, yeast extract, pH （7.25）, and temperature （28~C） conditions for the effective production of secondary metabolites. CONCLUSION： This study revealed that Streptomyces sp. VITPSA is an excellent source of secondary metabolites with various bioactivities.
