Medline/PubMed Citation | Related Articles in PubMed | Download to Citation Manager

Biotechnology and Applied Biochemistry (2002) 35, (149–154) (Printed in Great Britain)

PDF

Legacy HTML

Purification and characterization of alkaline protease from Alcaligenes faecalis

E. Berla Thangam and G. Suseela Rajkumar¹

Microbiology Division, Central Leather Research Institute, Adyar, Chennai 600 020, India

Key words: production, properties, serine protease.

Abbreviations used: BAPNA, benzoyl-D,L-arginine p-nitroanilide; BAEE, benzoyl-L-arginine ethyl ester.

¹To whom correspondence should be addressed (e-mail suseelarajkumar@hotmail.com).

Extracellular alkaline protease from the alkalophilic bacterium Alcaligenes faecalis was purified by a combination of ion-exchange and size-exclusion chromatographic methods, and its properties were examined. The purified enzyme had a specific activity of 563.8 µmol of tyrosine/min per mg of protein and gave a single band on native PAGE and SDS/PAGE with a molecular mass of 67 kDa. Gelatin zymogram also revealed one clear zone of proteolytic activity which corresponded to the band obtained with native PAGE and SDS/PAGE. The enzyme had an optimal pH of 9.0 and exhibited its highest activity at 55 °C. The enzyme activity was inhibited by PMSF, suggesting the presence of serine residues at the active site. The enzyme had a K_m of 1.66 mg/ml and a V_max of 526 units/min per mg of protein with casein as the substrate.

Received 2 March 2001/12 December 2001; accepted 15 January 2002

Portland Press Ltd © 2002