Journal:JBIC:26
From Proteopedia
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<StructureSection load='' size='450' side='right' scene='59/596313/Cv/15' caption=''> | <StructureSection load='' size='450' side='right' scene='59/596313/Cv/15' caption=''> | ||
=== Fluoride inhibition of Sporosarcina pasteurii urease: structure and thermodynamics === | === Fluoride inhibition of Sporosarcina pasteurii urease: structure and thermodynamics === | ||
| - | <big>Stefano Benini, Michele Cianci, Luca Mazzei and Stefano Ciurli</big> <ref> | + | <big>Stefano Benini, Michele Cianci, Luca Mazzei and Stefano Ciurli</big> <ref>PMID 25113581 </ref> |
<hr/> | <hr/> | ||
<b>Molecular Tour</b><br> | <b>Molecular Tour</b><br> | ||
| - | <scene name='59/596313/Cv/3'>Urease</scene> is a nickel-dependent enzyme (<span style="color:green;background-color:black;font-weight:bold;">Ni(II) ions are shown as green spheres</span>, <font color='darkmagenta'><b>α</b></font>, <span style="color:yellow;background-color:black;font-weight:bold;">β</span>, and <span style="color:deeppink;background-color:black;font-weight:bold;">γ</span> subunits are colored in <font color='darkmagenta'><b>darkmagenta</b></font>, <span style="color:yellow;background-color:black;font-weight:bold;">yellow</span>, <span style="color:deeppink;background-color:black;font-weight:bold;">deeppink</span>, respectively) and a virulence factor for ureolytic bacterial human pathogens, but it is also necessary to convert urea (see static image below), the most worldwide used fertiliser, into forms of nitrogen that can be taken up by crop plants. | + | <scene name='59/596313/Cv/3'>Urease</scene> (it is in homotrimeric form of αβγ heterotrimer) is a nickel-dependent enzyme (<span style="color:green;background-color:black;font-weight:bold;">Ni(II) ions are shown as green spheres</span>, <font color='darkmagenta'><b>α</b></font>, <span style="color:yellow;background-color:black;font-weight:bold;">β</span>, and <span style="color:deeppink;background-color:black;font-weight:bold;">γ</span> subunits are colored in <font color='darkmagenta'><b>darkmagenta</b></font>, <span style="color:yellow;background-color:black;font-weight:bold;">yellow</span>, <span style="color:deeppink;background-color:black;font-weight:bold;">deeppink</span>, respectively) and a virulence factor for ureolytic bacterial human pathogens, but it is also necessary to convert urea (see static image below), the most worldwide used fertiliser, into forms of nitrogen that can be taken up by crop plants. |
[[Image:Scheme_1.png|left|450px|thumb|]] | [[Image:Scheme_1.png|left|450px|thumb|]] | ||
{{Clear}} | {{Clear}} | ||
| - | 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 (<span style="color:gold;background-color:black;font-weight:bold;">both fluorides are colored in gold</span>). <scene name='59/596313/Cv/20'>Click here to see animation</scene>. |
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| + | Structural studies on ureases have revealed that the immediate environment around the two Ni(II) ions at the active site is conserved, as to induce a common mechanism of catalysis whose key step is the nucleophilic attack of the nickel-bridging hydroxide on the urea molecule bound to the bimetallic nickel cluster via O and N atoms (see static image below). | ||
| + | [[Image:Scheme_2.png|left|450px|thumb|]] | ||
| + | {{Clear}} | ||
| + | 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> (<span style="color:salmon;background-color:black;font-weight:bold;">colored in salmon</span>) 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> (<span style="color:cyan;background-color:black;font-weight:bold;">colored in cyan</span>) the Ni(II)-bridging hydroxide, blocking its nucleophilic attack on urea. | ||
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| + | '''PDB references:''' 1.58 A resolution native ''Sporosarcina pasteurii'' urease [[4ceu]]; 1.59 A resolution Fluoride inhibited ''Sporosarcina pasteurii'' urease [[4cex]]. | ||
</StructureSection> | </StructureSection> | ||
<references/> | <references/> | ||
__NOEDITSECTION__ | __NOEDITSECTION__ | ||
Current revision
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- ↑ Benini S, Cianci M, Mazzei L, Ciurli S. Fluoride inhibition of Sporosarcina pasteurii urease: structure and thermodynamics. J Biol Inorg Chem. 2014 Aug 12. PMID:25113581 doi:http://dx.doi.org/10.1007/s00775-014-1182-x
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