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      Production of Thermostable Organic Solvent Tolerant Keratinolytic Protease from Thermoactinomyces sp. RM4: IAA Production and Plant Growth Promotion

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          Abstract

          There are several reports about the optimization of protease production, but only few have optimized the production of organic solvent tolerant keratinolytic proteases that show remarkable exploitation in the development of the non-polluting processes in biotechnological industries. The present study was carried with aim to optimize the production of a thermostable organic solvent tolerant keratinolytic protease Thermoactinomyces sp. RM4 utilizing chicken feathers. Thermoactinomyces sp. RM4 isolated from the soil sample collected from a rice mill wasteyard site near Kashipur, Uttrakhand was identified on the basis of 16S rDNA analysis. The production of organic solvent tolerant keratinolytic protease enzyme by Thermoactinomyces sp. RM4 was optimized by varying physical culture conditions such as pH (10.0), temperature (60°C), inoculum percentage (2%), feather concentration (2%) and agitation rate (2 g) for feather degradation. The result showed that Thermoactinomyces sp. RM4 potentially produces extra-cellular thermostable organic solvent tolerant keratinolytic protease in the culture medium. Further, the feather hydrolysate from keratinase production media showed plant growth promoting activity by producing indole-3-acetic acid itself. The present findings suggest that keratinolytic protease from Thermoactinomyces sp. RM4 offers enormous industrial applications due to its organic solvent tolerant property in peptide synthesis, practical role in feather degradation and potential function in plant growth promoting activity, which might be a superior candidate to keep ecosystem healthy and functional.

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          Most cited references51

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          Rules for optimization of biocatalysis in organic solvents.

          General rules for the optimization of different biocatalytic systems in various types of media containing organic solvents are derived by combining data from the literature, and the logarithm of the partition coefficient, log P, as a quantitative measure of solvent polarity. (1) Biocatalysis in organic solvents is low in polar solvents having a log P 4. It was found that this correlation between polarity and activity parallels the ability of organic solvents to distort the essential water layer that stabilizes the biocatalysts. (2) Further optimization of biocatalysis in organic solvents is achieved when the polarity of the microenvironment of the biocatalyst (log P(i)) and the continuous organic phase (log P(cph)) is tuned to the polarities of both the substrate (log P(s)) and the product (log P(p)) according to the following rules: |log P(i) - log P(s)| and |log P(cph) - log P(p)| should be minimal and |log P(cph) - log P(s)| and |log P(i) - log P(p)| should be maximal, with the exception that in the case of substrate inhibition log P(i), should be optimized with respect to log P(s) In addition to these simple optimization rules, the future developments of biocatalysis in organic solvents are discussed.
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            CRYSTALLINE SOYBEAN TRYPSIN INHIBITOR

            M. Kunitz (1947)
            A study has been made of the general properties of crystalline soybean trypsin inhibitor. The soy inhibitor is a stable protein of the globulin type of a molecular weight of about 24,000. Its isoelectric point is at pH 4.5. It inhibits the proteolytic action approximately of an equal weight of crystalline trypsin by combining with trypsin to form a stable compound. Chymotrypsin is only slightly inhibited by soy inhibitor. The reaction between chymotrypsin and the soy inhibitor consists in the formation of a reversibly dissociable compound. The inhibitor has no effect on pepsin. The inhibiting action of the soybean inhibitor is associated with the native state of the protein molecule. Denaturation of the soy protein by heat or acid or alkali brings about a proportional decrease in its inhibiting action on trypsin. Reversal of denaturation results in a proportional gain in the inhibiting activity. Crystalline soy protein when denatured is readily digestible by pepsin, and less readily by chymotrypsin and by trypsin. Methods are given for measuring trypsin and inhibitor activity and also protein concentration with the aid of spectrophotometric density measurements at 280 mµ.
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              Microbial Enzymes: Tools for Biotechnological Processes

              Microbial enzymes are of great importance in the development of industrial bioprocesses. Current applications are focused on many different markets including pulp and paper, leather, detergents and textiles, pharmaceuticals, chemical, food and beverages, biofuels, animal feed and personal care, among others. Today there is a need for new, improved or/and more versatile enzymes in order to develop more novel, sustainable and economically competitive production processes. Microbial diversity and modern molecular techniques, such as metagenomics and genomics, are being used to discover new microbial enzymes whose catalytic properties can be improved/modified by different strategies based on rational, semi-rational and random directed evolution. Most industrial enzymes are recombinant forms produced in bacteria and fungi.
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                Author and article information

                Contributors
                Journal
                Front Microbiol
                Front Microbiol
                Front. Microbiol.
                Frontiers in Microbiology
                Frontiers Media S.A.
                1664-302X
                05 August 2016
                2016
                : 7
                : 1189
                Affiliations
                [1] 1Department of Biochemistry, G. B. Pant University of Agriculture and Technology Pantnagar, India
                [2] 2College of Basic Science & Humanities, Sardarkrushinagar Dantiwada Agricultural University Palanpur, India
                [3] 3Climate Change and Forest Influence Division, Forest Research Institute Dehradun, India
                [4] 4Department of Biotechnology, Bheemtal Campus, Kumaun University Nainital, India
                [5] 5Forest Pathology Division, Forest Research Institute Dehradun, India
                [6] 6Department of Botany, Zakir Husain Delhi College, University of Delhi New Delhi, India
                Author notes

                Edited by: Vijai Kumar Gupta, National University of Ireland, Galway, Ireland

                Reviewed by: Maulin P. Shah, Industrial Waste Water Research Lab, India; Ramesh Chandra Pant, G. B. Pant University of Agriculture and Technology, India

                *Correspondence: Amit Verma, amibiochem@ 123456rediffmail.com

                This article was submitted to Microbiotechnology, Ecotoxicology and Bioremediation, a section of the journal Frontiers in Microbiology

                Article
                10.3389/fmicb.2016.01189
                4974946
                27555836
                2d1cc7d6-d1c8-4038-ae25-41152e156432
                Copyright © 2016 Verma, Singh, Anwar, Kumar, Ansari and Agrawal.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                History
                : 06 June 2016
                : 19 July 2016
                Page count
                Figures: 7, Tables: 2, Equations: 0, References: 58, Pages: 13, Words: 0
                Funding
                Funded by: Department of Science and Technology, Ministry of Science and Technology 10.13039/501100001409
                Award ID: DST/017/2013
                Categories
                Microbiology
                Original Research

                Microbiology & Virology
                keratinase,thermoactinomyces,ecosystem health,feather degradation,organic solvent stability,indole-3-acetic acid production

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