Immobilization of pigeonpea (Cajanus cajan) urease on DEAE-cellulose paper strips for urea estimation.
Pigeonpea ( Cajanus cajan ) urease was immobilized on 1 cmx1 cm DEAE-cellulose paper strips. The optimum immobilization (51% activity) was observed at 4 degrees C, with a protein concentration of 1.0 mg/strip. The apparent optimum pH shifted from 7.3 to 6.8. Immobilized urease showed an optimal stability temperature of 67 degrees C, compared with 47 degrees C for the soluble urease. Time-dependent kinetics of the thermal inactivation of the immobilized urease were examined and found to be monophasic as compared with the soluble enzyme, which was biphasic. The Michaelis constant ( K (m)) for the DEAE-cellulose-immobilized urease was found to be 4.75 mM, 1.5 times higher than the soluble enzyme. Immobilized strips stored at 4 degrees C showed an increased half-life ( t (1/2)=150 days). There was practically no leaching of the enzyme from the immobilized strips over a period of 2 weeks. These strips were used for estimating the urea content of blood samples; the results obtained matched well with those obtained in a clinical laboratory through an Autoanalyzer(R) (Zydus Co., Rome, Italy). The easy availability of pigeonpea urease, the ease of its immobilization on DEAE-cellulose strips and the significantly lower cost of urease described in the present study makes it a suitable product for future applications in diagnostics.
Reddy K
Ravi Charan
RC
School of Biotechnology, Faculty of Science, Banaras Hindu University, Varanasi-221 005, India.
Srivastava
Punit K
PK
Dey
Prakash M
PM
Kayastha
Arvind M
AM
D036561:Cajanus / Q000201:enzymology*; D003636:DEAE-Cellulose / Q000737:chemistry*; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry*; D006207:Half-Life; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D010945:Plants, Edible / Q000201:enzymology*; D013696:Temperature; D013997:Time Factors; D014508:Urea / Q000097:blood*; D014510:Urease / Q000737:chemistry* / Q000302:isolation & purification
D004800:Enzymes, Immobilized; D014508:Urea; D003636:DEAE-Cellulose; D014510:Urease