Purification and characterization of the heat-stable serine proteinase from Thermomonospora fusca YX.
The proteinase secreted from Thermomonospora fusca YX grown on cellulose was purified by (NH4)2SO4 fractionation and cation-exchange chromatography. The isolated proteinase readily hydrolysed several proteins and demonstrated activity towards casein from 35 to 95 degrees C (at pH 8.0) with maximum activity at 80 degrees C. It exhibited broad pH and ionic-strength optima centered at pH 9.0 and 0.2 M-NaCl respectively, and it retained high activity in the presence of 2% (w/v) SDS, 20 mM-dithiothreitol and 1.0 M-NaCl. The proteinase, which was fully inhibited by phenylmethanesulphonyl fluoride, had an Mr of 14,500 and an isoelectric point at 9.21. A measurement of proteinase thermal stability demonstrated a T50% (15 min) of 85 degrees C at pH 4.5.
D000190:Actinomyces / Q000201:enzymology*; D002850:Chromatography, Gel; D004229:Dithiothreitol / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D008970:Molecular Weight; D009994:Osmolar Concentration; D010664:Phenylmethylsulfonyl Fluoride; D012697:Serine Endopeptidases / Q000302:isolation & purification*; D015842:Serine Proteinase Inhibitors; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D013696:Temperature
D015842:Serine Proteinase Inhibitors; D012967:Sodium Dodecyl Sulfate; D010664:Phenylmethylsulfonyl Fluoride; D012697:Serine Endopeptidases; D004229:Dithiothreitol