Extracellular acid protease from Rhizopus oryzae: purification and characterization

Kumar, Sushil ; Sharma, Neeru S. ; Saharan, Mukh R. ; Singh, Randhir (2005) Extracellular acid protease from Rhizopus oryzae: purification and characterization Process Biochemistry, 40 (5). pp. 1701-1705. ISSN 1359-5113

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Official URL: http://www.sciencedirect.com/science/article/pii/S...

Related URL: http://dx.doi.org/10.1016/j.procbio.2004.06.047


Extracellular aspartate protease from Rhizopus oryzae was purified 91 times with 26% recovery using (NH4)2SO4 fractionation, ion-exchange and size-exclusion chromatographic techniques. The enzyme was found to be monomeric in nature having a molecular mass of 34 kDa. The enzyme acts optimally at 60°C with activation energy of 15.16 kcal/mol and was stable in the temperature range of 30-45°C. The purified enzyme is an acid protease with optimum pH of 5.5 and retained 96% of residual activity between pH 5.5 to 7.5. Ca2+ activation (250 times) and varying substrate concentration gave an hyperbolic response. The Lineweaver-Burk plot showed Km value of 5 mg/ml, when skim milk was used as substrate. The enzyme inhibition of 73 and 93% by pepstatin at 10 and 20 μ M, respectively proved it to be an aspartate protease; however, the additional requirement of histidine residue for enzyme activity has been indicated by differential spectra of diethyl pyrocarbonate treated versus untreated enzyme.

Item Type:Article
Source:Copyright of this article belongs to Elsevier Science.
Keywords:Microbial Enzyme; Aspartate Protease; Milk Clotting Activity; Cheese; Rhizopus Oryzae
ID Code:50049
Deposited On:21 Jul 2011 14:44
Last Modified:21 Jul 2011 14:44

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