Full Name: rUrease (High Km)
Urea amidohydrolase (EC 220.127.116.11)
Host cell: E.coli
Specific activity: > 150 U/mg protein (at 25°C and pH 7.0)
Contaminant: (as rUrease activity = 100%)
NADPHox < 0.001 %
State: Lyophilized powder of highly purified enzyme. One mg of powder contains the following.
Protein 500 ± 300 μg
K-PO4 470 ± 300 μg
EDTA¥Na2 30 ± 10 μg
The Active mutant urease is expressed in E.Coli by recombinant DNA procedures and purified by conventional chromatographies. Mutant urease from Klebsiella aerogenes has a vast Km value (200mM).
The substitution of His219 at the active center of α subunit to other amino acid residues by protein engineering is known to modify the affinity against urea and to shift Km. Among those mutants, αH219Q has almost similar enzymatic properities except for a vast Km against urea. The Km value is 200mM, being 1,000 fold larger than that of wild type (0.2mM). The enzymatic properties of recombinant mutant urease (αH219Q) are summarized below.
1) Molecular weight: 224,000 (α 2 β 4 γ 4)
2) pH-activity profile: optimum around pH 7-8
3) pH-stability profile: stable in a range of pH4-10
4) Heat-activity profile: optimum around 50-65°C
5) Heat stability: less than 70°C
6) Km (urea): 200mM (pH 8)