Journal:JBIC:26
From Proteopedia
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| - | A strategy to control the activity of urease for medical and agricultural applications is to use enzyme inhibitors. Fluoride is a known urease inhibitor, but the structural basis of its mode of inhibition are still undetermined. Here, kinetic studies on the fluoride-induced inhibition of urease from ''Sporosarcina pasteurii'', a widespread and highly ureolytic soil bacterium, revealed a mixed competitive and uncompetitive mechanism. The pH-dependence of the inhibition constants, investigated in the 6.5-8.0 range, reveals a predominant uncompetitive mechanism that increases by increasing the pH, and a lesser competitive inhibition that increases by lowering the pH. Ten crystal structures of the enzyme were independently determined using five crystals of the <scene name='59/596313/Cv/13'>native form</scene> and five crystals of the protein crystallised in the presence of fluoride. The analysis of these structures revealed the presence of <scene name='59/596313/Cv/14'>two fluoride anions coordinated to the Ni(II) ions in the active site</scene>, in terminal and bridging positions. The present study consistently supports an interaction of fluoride with the nickel centres in the urease active site in which one fluoride competitively binds to the Ni(II) ion proposed to coordinate urea in the initial step of the catalytic mechanism, while another fluoride uncompetitively substitutes the Ni(II)-bridging hydroxide, blocking its nucleophilic attack on urea. | + | A strategy to control the activity of urease for medical and agricultural applications is to use enzyme inhibitors. Fluoride is a known urease inhibitor, but the structural basis of its mode of inhibition are still undetermined. Here, kinetic studies on the fluoride-induced inhibition of urease from ''Sporosarcina pasteurii'', a widespread and highly ureolytic soil bacterium, revealed a mixed competitive and uncompetitive mechanism. The pH-dependence of the inhibition constants, investigated in the 6.5-8.0 range, reveals a predominant uncompetitive mechanism that increases by increasing the pH, and a lesser competitive inhibition that increases by lowering the pH. Ten crystal structures of the enzyme were independently determined using five crystals of the <scene name='59/596313/Cv/13'>native form</scene> and five crystals of the protein crystallised in the presence of fluoride. The analysis of these structures revealed the presence of <scene name='59/596313/Cv/14'>two fluoride anions coordinated to the Ni(II) ions in the active site</scene>, in terminal and bridging positions. <scene name='59/596313/Cv/19'>Click here to see animation</scene>. The present study consistently supports an interaction of fluoride with the nickel centres in the urease active site in which <scene name='59/596313/Cv/17'>one fluoride competitively binds</scene> to the Ni(II) ion proposed to coordinate urea in the initial step of the catalytic mechanism, while <scene name='59/596313/Cv/18'>another fluoride uncompetitively substitutes</scene> the Ni(II)-bridging hydroxide, blocking its nucleophilic attack on urea. |
</StructureSection> | </StructureSection> | ||
<references/> | <references/> | ||
__NOEDITSECTION__ | __NOEDITSECTION__ | ||
Revision as of 13:02, 4 August 2014
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- ↑ REF
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