User:Judy Voet/Lysozyme
From Proteopedia
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| - | <applet load='aln_1H6M_to_1HEW_2.pdb' size='300' frame='true' align='right' caption=' | + | <applet load='aln_1H6M_to_1HEW_2.pdb' size='300' frame='true' align='right' caption=' Hen Egg White (HEW) Lysozyme containing a trisaccharide of N-acetylglucosamine (NAG) bound to the active site, PDBid 1HEW' scene='User:Judy_Voet/Lysozyme/Lysozyme1/12' /> |
Lysozyme was the first enzyme whose x-ray structure was determined <ref> PMID 5840126</ref><ref>Phillips, D. C. The hen egg white lysozyme molecule. Proc. Natl Acad. Sci. USA 57, 483-495 (1967)</ref>. This scene shows Hen Egg White (HEW) Lysozyme containing a trisaccharide of N-acetylglucosamine (NAG) bound to the active site. David Phillips, who determined the structure in 1965, saw that the active site was large enough to fit three more saccharide units and his group built a model extending the trisaccharide to a hexasaccharide | Lysozyme was the first enzyme whose x-ray structure was determined <ref> PMID 5840126</ref><ref>Phillips, D. C. The hen egg white lysozyme molecule. Proc. Natl Acad. Sci. USA 57, 483-495 (1967)</ref>. This scene shows Hen Egg White (HEW) Lysozyme containing a trisaccharide of N-acetylglucosamine (NAG) bound to the active site. David Phillips, who determined the structure in 1965, saw that the active site was large enough to fit three more saccharide units and his group built a model extending the trisaccharide to a hexasaccharide | ||
<scene name='User:Judy_Voet/Lysozyme/Lysozyme1_hexamer/1'>HEW Lysozyme with hexamer</scene> fit into the active site <ref> coordinates of the model kindly provided by Louise Johnson</ref>. Alternately click on <scene name='User:Judy_Voet/Lysozyme/Lysozyme1/15'>trimer</scene> and <scene name='User:Judy_Voet/Lysozyme/Lysozyme1_hexamer/1'>hexamer</scene> to turn the model portion of the hexasaccharide on and off. | <scene name='User:Judy_Voet/Lysozyme/Lysozyme1_hexamer/1'>HEW Lysozyme with hexamer</scene> fit into the active site <ref> coordinates of the model kindly provided by Louise Johnson</ref>. Alternately click on <scene name='User:Judy_Voet/Lysozyme/Lysozyme1/15'>trimer</scene> and <scene name='User:Judy_Voet/Lysozyme/Lysozyme1_hexamer/1'>hexamer</scene> to turn the model portion of the hexasaccharide on and off. | ||
The interesting thing about the model-building experiment was that the only way that the hexasaccharide would fit into the active site was for the 4th saccharide to be strained into a <scene name='User:Judy_Voet/Lysozyme/Lysozyme1/5'>half-chair conformation</scene>. This conformation is what would be necessary for the formation of an oxocarbenium ion (oxionium ion). When the model was studied, <scene name='User:Judy_Voet/Lysozyme/Lysozyme1/8'>Glu 35</scene> was found to be in an ideal location to act as a general acid catalyst, protonating bridging oxygen between the 4th and 5th saccharide units. <scene name='User:Judy_Voet/Lysozyme/Lysozyme1/14'>Asp 52</scene> was too far away in the static lysozyme structure to have formed a covalent bond with C1 of the half-chair hexasaccharide 4, so Phillips proposed that it acted as an electrostatic stabilizer of the oxonium ion (referred to as The Phillips Mechanism). | The interesting thing about the model-building experiment was that the only way that the hexasaccharide would fit into the active site was for the 4th saccharide to be strained into a <scene name='User:Judy_Voet/Lysozyme/Lysozyme1/5'>half-chair conformation</scene>. This conformation is what would be necessary for the formation of an oxocarbenium ion (oxionium ion). When the model was studied, <scene name='User:Judy_Voet/Lysozyme/Lysozyme1/8'>Glu 35</scene> was found to be in an ideal location to act as a general acid catalyst, protonating bridging oxygen between the 4th and 5th saccharide units. <scene name='User:Judy_Voet/Lysozyme/Lysozyme1/14'>Asp 52</scene> was too far away in the static lysozyme structure to have formed a covalent bond with C1 of the half-chair hexasaccharide 4, so Phillips proposed that it acted as an electrostatic stabilizer of the oxonium ion (referred to as The Phillips Mechanism). | ||
| - | <applet load='aln_1H6M_to_1HEW_2.pdb' size='300' frame='true' align='right' caption=' | + | <applet load='aln_1H6M_to_1HEW_2.pdb' size='300' frame='true' align='right' caption='NAG-2-fluoro-glucosyl fluoride (NAG2FGlcF) bound to Glu35Gln HEW Lysozyme PDBid 1H6M' scene='User:Judy_Voet/Lysozyme/1h6m/3'/> |
Then, in 2001, Stephen Withers published <scene name='User:Judy_Voet/Lysozyme/1h6m/3'>1H6M</scene>,<ref>PMID 11518970</ref> in which Glu 35 was mutated to Gln to remove general acid catalysis and the substrate contained NAG-2-fluoro-glucosyl fluoride (NAG2FGlcF). The fluoro group does not require acid catalysis to be a good leaving group, and the remaining saccharide, in the absence of the acid necessary to catalyse the second step of the reaction, was demonstrated to <scene name='User:Judy_Voet/Lysozyme/1h6m/4'>form a covalent intermediate</scene>. Note in this <scene name='User:Judy_Voet/Lysozyme/1h6m/6'>superposition</scene> of half chair model with 1HEW (greens) on covalent intermediate in 1H6M (blues) the motion of Asp 52 and C1 of the sugar ring in going from the model to the covalent intermediate. | Then, in 2001, Stephen Withers published <scene name='User:Judy_Voet/Lysozyme/1h6m/3'>1H6M</scene>,<ref>PMID 11518970</ref> in which Glu 35 was mutated to Gln to remove general acid catalysis and the substrate contained NAG-2-fluoro-glucosyl fluoride (NAG2FGlcF). The fluoro group does not require acid catalysis to be a good leaving group, and the remaining saccharide, in the absence of the acid necessary to catalyse the second step of the reaction, was demonstrated to <scene name='User:Judy_Voet/Lysozyme/1h6m/4'>form a covalent intermediate</scene>. Note in this <scene name='User:Judy_Voet/Lysozyme/1h6m/6'>superposition</scene> of half chair model with 1HEW (greens) on covalent intermediate in 1H6M (blues) the motion of Asp 52 and C1 of the sugar ring in going from the model to the covalent intermediate. | ||
===Some Useful External Links=== | ===Some Useful External Links=== | ||
Revision as of 15:48, 4 November 2009
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Lysozyme was the first enzyme whose x-ray structure was determined [1][2]. This scene shows Hen Egg White (HEW) Lysozyme containing a trisaccharide of N-acetylglucosamine (NAG) bound to the active site. David Phillips, who determined the structure in 1965, saw that the active site was large enough to fit three more saccharide units and his group built a model extending the trisaccharide to a hexasaccharide fit into the active site [3]. Alternately click on and to turn the model portion of the hexasaccharide on and off. The interesting thing about the model-building experiment was that the only way that the hexasaccharide would fit into the active site was for the 4th saccharide to be strained into a . This conformation is what would be necessary for the formation of an oxocarbenium ion (oxionium ion). When the model was studied, was found to be in an ideal location to act as a general acid catalyst, protonating bridging oxygen between the 4th and 5th saccharide units. was too far away in the static lysozyme structure to have formed a covalent bond with C1 of the half-chair hexasaccharide 4, so Phillips proposed that it acted as an electrostatic stabilizer of the oxonium ion (referred to as The Phillips Mechanism).
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Then, in 2001, Stephen Withers published ,[4] in which Glu 35 was mutated to Gln to remove general acid catalysis and the substrate contained NAG-2-fluoro-glucosyl fluoride (NAG2FGlcF). The fluoro group does not require acid catalysis to be a good leaving group, and the remaining saccharide, in the absence of the acid necessary to catalyse the second step of the reaction, was demonstrated to . Note in this of half chair model with 1HEW (greens) on covalent intermediate in 1H6M (blues) the motion of Asp 52 and C1 of the sugar ring in going from the model to the covalent intermediate.
Some Useful External Links
Retaining Glycoside Hydrolases
References
- ↑ Johnson LN, Phillips DC. Structure of some crystalline lysozyme-inhibitor complexes determined by X-ray analysis at 6 Angstrom resolution. Nature. 1965 May 22;206(986):761-3. PMID:5840126
- ↑ Phillips, D. C. The hen egg white lysozyme molecule. Proc. Natl Acad. Sci. USA 57, 483-495 (1967)
- ↑ coordinates of the model kindly provided by Louise Johnson
- ↑ Vocadlo DJ, Davies GJ, Laine R, Withers SG. Catalysis by hen egg-white lysozyme proceeds via a covalent intermediate. Nature. 2001 Aug 23;412(6849):835-8. PMID:11518970 doi:10.1038/35090602
