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Extracellular acid protease from Aspergillus niger
I1: purification and characterization
Rayda Siala#,
Alya Sellami-Kamoun#, Mohamed Hajji, Ines Abid,
Neji Gharsallah and Moncef Nasri*
Laboratoire de Génie Enzymatique et de Microbiologie, Ecole
Nationale d'Ingénieurs de Sfax, B.P. (W) 3038 Sfax, Tunisia.
*Corresponding author.
E-mail:
Moncef.Nasri@enis.rnu.tn or
mon_nasri@yahoo.fr.
Tel: +216 74 274 088. Fax: +216 74 275 595.
#These authors contribute equally to this article
Accepted 30 April, 2009 |
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A new strain of Aspergillus niger producing acid
protease was isolated and identified by universal primers
NL1 and NL4. The acid protease from A. niger I1 was
purified to homogeneity by ultrafiltration using a 10-KDa
cut-off membrane, gel filtration on Sephadex G-75 and ion
exchange chromatography on CM-Sephadex C-50, with a
3.55-fold increase in specific activity and 56% recovery.
The molecular weight of the protease was estimated to be 50
kDa on SDS-PAGE and gel filtration, which is higher than
those from other A. niger strains. Carbohydrate
content of the purified protease, determined by the chemical
anthrone method, was calculated to be 16%. The Km and
Vmax for caseinolytic activity of the purified enzyme were
found to be 1.02 mM and 2.2 µmol/min, respectively. The
enzyme was optimally active at 60°C and pH 3.0. The most
metal ions tested had no significant effect on protease
activity. The enzyme activity was inhibited by pepstatin A,
suggesting that the purified enzyme is an aspartic protease.
Key words:
Acid protease, Aspergillus niger, purification,
aspergillopepsin, glycosylation. |
