title: Isolation and characterization of a metal ion-dependent alkaline protease from a halotolerant Bacillus aquimaris VITP4.
abstract: A halotolerant bacterium Bacillus acquimaris VITP4 was used for the production of extracellular protease. Fractional precipitation using ammonium chloride was used to obtain the enzyme. The protease exhibited optimum activity at pH 8.0 and 40 degrees C and retained 50% of its optimal proteolytic activity even in the presence of 4 M NaCl, suggesting that it is halotolerant. The molecular mass of protease, as revealed by SDS-PAGE was found to be 34 kDa and the homogeneity of the enzyme was confirmed by gelatin zymography and reverse-phase HPLC. Upon purification, the specific activity of th enzyme increased from 533 U/mg to 1719 U/mg. Protease inhibitors like phenyl methane sulphonyl fluoride and 2-mercaptoethanol did not affect the activity of the enzyme, but EDTA inhibited the activity, indicating the requirement of metal ions for activity. Cu2, Ni2+ and Mn2+ enhanced the enzyme activity, but Zn2+, Hg2+ and Fe2+ decreased the activity, while Mg2+, Ca2+ and K+ had no effect on the enzyme activity. The protease was quite stable in the presence of cationic (CTAB), anionic (SDS) and neutral detergents (Triton X-100 and Tween-20) and exhibited antimicrobial activity against selected bacterial and fungal strains. The stability characteristics and broad spectrum antimicrobial activity indicated the potential use of this protease in industrial applications.
authors.affiliation: School of Biosciences and Technology, VIT University, Vellore 632014, India.
authors.forename: Pooja
authors.initials: P
authors.lastname: Shivanand
authors.forename: Gurunathan
authors.initials: G
authors.lastname: Jayaraman
mesh_terms: D000890:Anti-Infective Agents / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology*; D001407:Bacillus / Q000145:classification / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid; D003902:Detergents / Q000494:pharmacology; D004586:Electrophoresis; D010450:Endopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology*; D004795:Enzyme Stability / Q000187:drug effects; D005110:Extracellular Space / Q000201:enzymology; D005658:Fungi / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000494:pharmacology*; D011480:Protease Inhibitors / Q000494:pharmacology; D012965:Sodium Chloride / Q000494:pharmacology; D013696:Temperature
chemical_list: D000890:Anti-Infective Agents; D001426:Bacterial Proteins; D003902:Detergents; D008670:Metals; D011480:Protease Inhibitors; D012965:Sodium Chloride; D010450:Endopeptidases; C087822:alkaline protease