title: Purification, characterization and thermodynamics of antifungal protease from Streptomyces sp. A6.
abstract: A 20 kDa antifungal serine protease from Streptomyces sp. A6 was purified to 34.56 folds by gel permeation chromatography. The enzyme exhibited highest activity at neutral to near alka- line pH 7-9 and 55 °C. Neutral surfactant triton X-100 enhanced the activity by 4.12 fold. The protease activity also increased (109.9-119%) with increasing concentration of urea (2-8 mole/l). The enzyme was identified as serine protease with 67% similarity to SFase 2 of Streptomyces fradiae by MALDI-LC-MS/MS analysis. Determination of kinetic constants k(m) , V(max) , k(cat) and k(cat) /k(m) suggested higher affinity of enzyme for N-Suc-Ala-Ala-Val-Ala-p NA (synthetic substrate for chymotrypsin activity). The enzyme was highly stable at temperature prevailing under field conditions (40 °C) as apparent from K(d) and t(1/2) values, 0.0065 and 106.75 min, respectively and high ΔG* and negative ΔS * values, 87.17 KJ/mole and -126.95 J/mole, respectively. Thermal stability and increased activity of protease in presence of commonly used chemical fertilizer, urea, suggested its feasibility for agricultural applications. The present study is the first report on thermodynamic and kinetic properties of an antifungal protease from Streptomyces sp. A6. The study reflects potential of this enzyme for biocontrol of fungal plant pathogens.
authors.affiliation: Department of Microbiology and Biotechnology Centre, Faculty of Science, The Maharaja Sayajirao University of Baroda, Vadodara, India.
authors.forename: Anil Kumar
authors.initials: AK
authors.lastname: Singh
authors.forename: Hari S
authors.initials: HS
authors.lastname: Chhatpar
mesh_terms: D000935:Antifungal Agents / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D012697:Serine Endopeptidases / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D013302:Streptomyces / Q000145:classification / Q000201:enzymology* / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D000935:Antifungal Agents; D015842:Serine Proteinase Inhibitors; D012697:Serine Endopeptidases