Pseudomonas aeruginosa A2 elastase: purification, characterization and biotechnological applications.
An extracellular protease from Pseudomonas aeruginosa A2 grown in media containing shrimp shell powder as a unique source of nutriments was purified and characterized. The enzyme was purified to homogeneity from culture supernatant by ultrafiltration, Sephadex G-100 gel filtration and Sepharose Mono Q anion exchange chromatography, with a 2.23-fold increase in specific activity and 64.3% recovery. The molecular mass of the enzyme was estimated to be 34 kDa. Temperature and pH with highest activity were 60 °C and 8.0, respectively. The protease activity was inhibited by EDTA suggesting that the purified enzyme is a metalloprotease. The enzyme is stable in the presence of organic solvents mainly diethyl ether and DMSO. The lasB gene, encoding the A2 elastase, was isolated and its DNA sequence was determined. The A2 protease was tested for shrimp waste deproteinization in the process of chitin preparation. The percent of protein removal after 3 h hydrolysis at 40 °C with an enzyme/substrate (E/S) ratio of 5 U/mg protein was about 75%. Additionally, A2 proteolytic preparation demonstrated powerful depilating capabilities of hair removal from bovine skin. Considering its promising properties, P. aeruginosa A2 protease may be considered a potential candidate for future use in several biotechnological processes.
Laboratoire de Génie Enzymatique et de Microbiologie, Université de Sfax, Ecole Nationale d'Ingénieurs de Sfax, B.P.1173 - 3038 Sfax, Tunisia. olfa ghor@yahoo.fr
Olfa
O
Ghorbel-Bellaaj
Ben Khaled
BK
Hayet
Ahmed
A
Bayoudh
Islem
I
Younes
Noomen
N
Hmidet
Kemel
K
Jellouli
Moncef
M
Nasri
D000595:Amino Acid Sequence; D060105:Animal Shells / Q000737:chemistry; D000818:Animals; D001709:Biotechnology / Q000379:methods*; D002417:Cattle; D002686:Chitin / Q000737:chemistry; D004795:Enzyme Stability; D006204:Hair Removal; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000494:pharmacology; D008969:Molecular Sequence Data; D009930:Organic Chemicals / Q000494:pharmacology; D010196:Pancreatic Elastase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D011550:Pseudomonas aeruginosa / Q000201:enzymology* / Q000235:genetics; D017421:Sequence Analysis; D012997:Solvents / Q000494:pharmacology; D013696:Temperature
D008670:Metals; D009930:Organic Chemicals; D012997:Solvents; D002686:Chitin; D010196:Pancreatic Elastase