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  • Purification of urease from jack bean (Canavalia ensiformis) with copper (II) chelated poly(hydroxyethyl methacrylate-N-methacryloyl-(L)-histidine methyl ester) cryogels.

Purification of urease from jack bean (Canavalia ensiformis) with copper (II) chelated poly(hydroxyethyl methacrylate-N-methacryloyl-(L)-histidine methyl ester) cryogels.

Journal of molecular recognition : JMR (2012-10-31)
Pınar Tekiner, Işik Perçin, Bahar Ergün, Handan Yavuz, Erol Aksöz
ABSTRACT

Jack bean (Canavalia ensiformis) is the source of interesting proteins that contribute to modern biochemistry, and urease is the primary of these proteins. Owing to its role and occurrence in nature, urease has become a part of extensive studies. In this study, jack bean urease (JBU) was purified by immobilized metal affinity chromatography using Cu(2+) chelated poly(hydroxyethyl methacrylate-N-methacryloyl-(L)-histidine methyl ester) [PHEMAH-Cu(2+)]-based cryogels. PHEMAH-Cu(2+) cryogel was synthesized and characterized for swelling degree, morphology (by SEM), N-methacryloyl-(L)-histidine methyl ester and Cu(2+) incorporation (by elemental analysis and atomic absorption spectrophotometry). The binding of JBU to PHEMAH-Cu(2+) cryogel was optimized by examining the effect of pH, flow rate and JBU concentration on binding. The maximal binding of JBU was 23.2 mg/dry gram of adsorbent. The maximal binding of JBU extracted from jack bean meal was 67.8 mg/dry gram of adsorbent. The elution of JBU from cryogel column was accomplished by 1.0 M NaCl in 20 mM phosphate buffer (pH 8.0). Molecular weight and purity of JBU from jack bean meal was estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It was observed that JBU could be repeatedly bound and eluted from (PHEMAH)-Cu(2+) cryogel with less than 10% loss in column capacity.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Urease from Canavalia ensiformis (Jack bean), Type III, powder, 15,000-50,000 units/g solid
Sigma-Aldrich
Urease from Canavalia ensiformis (Jack bean), Type C-3, powder, ≥600,000 units/g solid
Sigma-Aldrich
Urease from Canavalia ensiformis (Jack bean)
Sigma-Aldrich
Urease from Canavalia ensiformis (Jack bean), powder, ~1 U/mg
Sigma-Aldrich
Urease from Canavalia ensiformis (Jack bean), Type IX, powder, 50,000-100,000 units/g solid
Sigma-Aldrich
Urease from Canavalia ensiformis (Jack bean), powder, white, ~8 U/mg
Sigma-Aldrich
Urease from Canavalia ensiformis (Jack bean), Type III, glycerol solution, 500-800 units/mL