Urease isoelectric point

The isoelectric point, determined by Sumner and Hand (50) to have a value of 5.0-5.1, has been redetermined by the electrofocusing technique (43). The value obtained was 4.8. The solubility is extremely small at this pH, but the urease can be located by its enzymic action. It is a point of interest that the solubility of the isoelectric species increases spectacularly if the sulfhydryl groups are substituted with A-ethylmaleimide (NEM) (43). 

Urease has a molecular weight of590 000 30 000 and consists of six identical subunits. Each subunit contains two Ni ions of different valency which are involved in substrate binding and conversion. The isoelectric point of the protein is at pH 5 and the temperature optimum of the catalysis at 60°C. The kinetic constants for urea hydrolysis have been determined to be k+2 = 5870 s"1 and Km = 2.9 mmol/1. Other amides, such as formamide and semicarbazide, react much more slowly than urea. The pH optimum of urease depends on the nature of the buffer used and, with the exception of acetate buffer, equals the pJTs value of the buffer. The active center of urease contains an SH-group that is essential for the stability of the enzyme. Complexing agents, such as EDTA and reductants, are required for stabilization. 

The proteins ovalbumin (isoelectric point pi = 4.6), urease (pi = 5.0), and myoglobin (pi = 7.0) were added to a column of the anion-exchange resin, DEAE-cellulose at pH 6.5. The column was eluted with a dilute pH 6.5 buffer and then with the same buffer as a gradient, containing an increasing concentration of NaCl. In what order will the proteins be eluted from the column ... 

Urease is a simple protein with a molecular weight of 473,000 and an isoelectric point of 5.0-5.1. It contains 31 sulfhydryl groups per mole, not all of which are required for activity. Urease is sensitive, however, to both heavy metals and oxidizing agents. 

---