Papers by ASHWINI PUNTAMBEKAR
Research Journal of Biotechnology, Jun 25, 2022
Research Journal of Biotechnology
The use of enzymes in applied biotechnology has progressively increased in both industrial proces... more The use of enzymes in applied biotechnology has progressively increased in both industrial processes, products and in medical field. Proteolytic enzymes play an important regulatory role in many physiological processes and also represent a therapeutic target for several diseases including cancer, hypertension, blood clotting disorders, respiratory and viral infection. Proteases, a largest and ubiquitous class of enzymes, have a divergent role in biomedical field. The current review includes the basic information about the protease classification and optimized growth parameters to maximize the production of alkaline proteases and applications of proteases in a wide variety of industries including leather, textile, food manufacturing, pharmaceutical, detergent and waste management. The review also implicates the importance of genetic tools to obtain the novel engineered protease with improved catalytic performance and stability, pH and thermal tolerance.
Asian Journal of Pharmaceutical and Clinical Research, 2017
Objective: The main objective of this study is to investigate the industrial applications of a ... more Objective: The main objective of this study is to investigate the industrial applications of a thermophillic alkaline protease from a hot water spring bacterial isolate “A” and to study its production, optimization, and purification.Methods: The alkaline protease was produced using shake flask studies maintaining a pH of 9.0 and a temperature of 50°C. Optimization studies of the enzyme were carried out using variable pH, temperature, organic carbon, and nitrogen sources followed by purification of the enzyme using DEAE-cellulose ion exchange chromatography technique. Stability of the enzyme was analyzed in the presence of organic solvents and surfactants. The efficiency of the enzyme in the removal of proteinaceous stains in the presence of strong detergents under extreme conditions was assessed. The fibrinolytic activity of the enzyme in dissolving the blood clot was confirmed.Results: The isolated alkaline protease was purified to homogeneity with a 16-fold increase. Media optim...
International Journal of Pharmacy and Pharmaceutical Sciences, 2017
Objective: The main objective of the study was isolation, purification and characterization of pr... more Objective: The main objective of the study was isolation, purification and characterization of protein protease inhibitor from the seeds of Phaseolus vulgaris and analysis of its antimicrobial potential.Methods: The protease inhibitor was extracted by homogenizing seeds of Phaseolus vulgaris in 0.1 M phosphate buffer (pH-7.0). The crude extract of the inhibitor was purified by using ammonium sulphate precipitation followed by DEAE Cellulose ion exchange chromatography. The protease inhibitor was characterized to determine its optimum pH and pH stability, optimum temperature and temperature stability, stability in the presence of chemical modifiers, thermal stabilizers, metal ions, detergents, oxidising and reducing agents. The antimicrobial potential of the inhibitor against various bacterial species was confirmed using agar well diffusion method.Results: The extracted protease inhibitor was purified to homogeneity with a 1.4 fold increase in the specific activity and 56 % purificat...
International Journal of Pharmacy and Pharmaceutical Sciences, Sep 1, 2017
Objective: The main objective of the study was isolation, purification and characterization of pr... more Objective: The main objective of the study was isolation, purification and characterization of protein protease inhibitor from the seeds of Phaseolus vulgaris and analysis of its antimicrobial potential. Methods: The protease inhibitor was extracted by homogenizing seeds of Phaseolus vulgaris in 0.1 M phosphate buffer (pH-7.0). The crude extract of the inhibitor was purified by using ammonium sulphate precipitation followed by DEAE Cellulose ion exchange chromatography. The protease inhibitor was characterized to determine its optimum pH and pH stability, optimum temperature and temperature stability, stability in the presence of chemical modifiers, thermal stabilizers, metal ions, detergents, oxidising and reducing agents. The antimicrobial potential of the inhibitor against various bacterial species was confirmed using agar well diffusion method. Results: The extracted protease inhibitor was purified to homogeneity with a 1.4 fold increase in the specific activity and 56 % purification yield. Inhibitor was optimally active at pH 7.0 and a temperature of 50 °C as well as showed considerable stability over pH ranging from 4.0-11.0 and up to a temperature of 70 °C for 4 h duration. The inhibitor was substantially active and stable in the presence of surfactants 1 % (v/v) Tween 20, 4 % (v/v) of oxidizing agents such as dimethyl sulphoxide (DMSO) and hydrogen peroxide (H2O2), 0.8 % (v/v) of reducing agents βmercaptoethanol and Sodium thioglycolate. Metal ions Mg ++ , Ca ++ and Zn ++ enhanced the activity of inhibitor while CaCl2, glycine and glycerol promoted thermal stability of the inhibitor. Chemical modification of amino acids at the active site by diethyl pyrocarbonate (DEPC) and phenyl methyl sulphonyl fluoride (PMSF) led to decrease in the inhibitory activity. The stoichiometry of trysin-protease inhibitor interaction was 1:2 while 216 μg of the inhibitor effected 50 % inhibition. The inhibitor displayed antimicrobial activity against Aeromonas hydrophilla, Citrobacter freundii and Acinetobacter baumanii. Conclusion: The experimental results confirmed the anti proteolytic action of protease inhibitor extracted from P. vulgaris as well as its promising antimicrobial properties. Thus isolated protease inhibitor has significant industrial and therapeutic potential.
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Papers by ASHWINI PUNTAMBEKAR