Urease location

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). 

M Polarization of the carbonyl bond by coordination activates it for nucleophilic attack. This situation is assumed to occur by the bridging hydroxide, which appears to be most suitably arranged when the picolinamide-0 is located in a bridging position, although its transient shift to a terminal site cannot be ruled out. The failure of 32 to hydrolyze picolinamide confirms that generation of a nucleophilic hydroxide is required for the reaction to proceed (82). In subsequent work, 33 was then found to serve as a valuable functional model for the urease active site (see Section III.B.4). 

Fig. 8.4. Biosensors may become interesting for probe detection. In this example (from Olson et al., 1991), a biotinylated and a fluoresceinated probe are hybridized in solution to the target (I). The primer sequences should be located outside this region for PCR-product detection. Avidin acts as a bridge between the biotinylated probe and the biotinylated surface of the biosensor (II). Fluorescein is detected with a urease-antibody conjugate. The rise in pH, after the addition of urea, is detected with the pH-sensitive biosensor. The total assay time is less than 2 h with a detectability of 30 amol and a coefficient of variation (standard deviation/mean) of less than 10%.

Fig. 1. Ribbon representation of an afiy unit of the crystallographic trimer of K. aerogeties urease. The Ni ions are located at the top of the a subunit (light gray) and are accessible from the solvent medium. The / and y subunits are depicted in dark and medium grey respectively. This drawing was prepared with Molscript [126] by A. Volbeda. Coordinates were obtained from entry 1KAU [16] from the Protein Data Bank [127]...

The key elements of urease s active she are two ruckel ions and a molecule of water located inside a structure vdiich creates a trap for a molecule of urea. A potent inhibitor can block this site or inactivate it by deforming its geometry. Some of the known compounds vdiich can thus save urea fiom die en matic destruction have been used so fiu in various areas such as medicine [1,2], agricuhute and animal fiuming [3]. 

BURNS R.G., PUKITE A.H. and McLAREN A.D. 1972. Concerning the location and persistence of soil urease. Soil Science Society of America Proceedings, 308-311. 

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