1rcj
From Proteopedia
(New page: 200px<br /><applet load="1rcj" size="450" color="white" frame="true" align="right" spinBox="true" caption="1rcj, resolution 1.63Å" /> '''Crystal structure of...) |
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| - | [[Image:1rcj.jpg|left|200px]]<br /><applet load="1rcj" size=" | + | [[Image:1rcj.jpg|left|200px]]<br /><applet load="1rcj" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1rcj, resolution 1.63Å" /> | caption="1rcj, resolution 1.63Å" /> | ||
'''Crystal structure of E166A mutant of SHV-1 beta-lactamase with the trans-enamine intermediate of tazobactam'''<br /> | '''Crystal structure of E166A mutant of SHV-1 beta-lactamase with the trans-enamine intermediate of tazobactam'''<br /> | ||
==Overview== | ==Overview== | ||
| - | Many pathogenic bacteria develop antibiotic resistance by utilizing | + | Many pathogenic bacteria develop antibiotic resistance by utilizing beta-lactamases to degrade penicillin-like antibiotics. A commonly prescribed mechanism-based inhibitor of beta-lactamases is tazobactam, which can function either irreversibly or in a transient manner. We have demonstrated previously that the reaction between tazobactam and a deacylation deficient variant of SHV-1 beta-lactamase, E166A, could be followed in single crystals using Raman microscopy [Helfand, M. S., et al. (2003) Biochemistry 42, 13386-13392]. The Raman data show that maximal populations of an enamine-like intermediate occur 20-30 min after "soaking in" has commenced. By flash-freezing crystals in this time frame, we were able to trap the enamine species. The resulting 1.63 A resolution crystal structure revealed tazobactam covalently bound in the trans-enamine intermediate state with close to 100% occupancy in the active site. The Raman data also indicated that tazobactam forms a larger population of enamine than sulbactam or clavulanic acid does and that tazobactam's intermediate is also the most long-lived. The crystal structure provides a rationale for this finding since only tazobactam is able to form favorable intra- and intermolecular interactions in the active site that stabilize this trans-enamine intermediate. These interactions involve both the sulfone and triazolyl groups that distinguish tazobactam from clavulanic acid and sulbactam, respectively. The observed stabilization of the transient intermediate of tazobactam is thought to contribute to tazobactam's superior in vitro and in vivo clinical efficacy. Understanding the structural details of differing inhibitor effectiveness can aid the design of improved mechanism-based beta-lactamase inhibitors. |
==About this Structure== | ==About this Structure== | ||
| - | 1RCJ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Klebsiella_pneumoniae Klebsiella pneumoniae] with MA4 and TBI as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Beta-lactamase Beta-lactamase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.5.2.6 3.5.2.6] Full crystallographic information is available from [http:// | + | 1RCJ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Klebsiella_pneumoniae Klebsiella pneumoniae] with <scene name='pdbligand=MA4:'>MA4</scene> and <scene name='pdbligand=TBI:'>TBI</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Beta-lactamase Beta-lactamase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.5.2.6 3.5.2.6] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1RCJ OCA]. |
==Reference== | ==Reference== | ||
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[[Category: Klebsiella pneumoniae]] | [[Category: Klebsiella pneumoniae]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
| - | [[Category: Akker, F | + | [[Category: Akker, F van den.]] |
| - | [[Category: Bonomo, R | + | [[Category: Bonomo, R A.]] |
| - | [[Category: Carey, M | + | [[Category: Carey, M P.]] |
| - | [[Category: Carey, P | + | [[Category: Carey, P R.]] |
| - | [[Category: Helfand, M | + | [[Category: Helfand, M S.]] |
| - | [[Category: Hujer, A | + | [[Category: Hujer, A M.]] |
| - | [[Category: Padayatti, P | + | [[Category: Padayatti, P S.]] |
| - | [[Category: Totir, M | + | [[Category: Totir, M A.]] |
[[Category: MA4]] | [[Category: MA4]] | ||
[[Category: TBI]] | [[Category: TBI]] | ||
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[[Category: penicillinase]] | [[Category: penicillinase]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 14:49:25 2008'' |
Revision as of 12:49, 21 February 2008
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Crystal structure of E166A mutant of SHV-1 beta-lactamase with the trans-enamine intermediate of tazobactam
Overview
Many pathogenic bacteria develop antibiotic resistance by utilizing beta-lactamases to degrade penicillin-like antibiotics. A commonly prescribed mechanism-based inhibitor of beta-lactamases is tazobactam, which can function either irreversibly or in a transient manner. We have demonstrated previously that the reaction between tazobactam and a deacylation deficient variant of SHV-1 beta-lactamase, E166A, could be followed in single crystals using Raman microscopy [Helfand, M. S., et al. (2003) Biochemistry 42, 13386-13392]. The Raman data show that maximal populations of an enamine-like intermediate occur 20-30 min after "soaking in" has commenced. By flash-freezing crystals in this time frame, we were able to trap the enamine species. The resulting 1.63 A resolution crystal structure revealed tazobactam covalently bound in the trans-enamine intermediate state with close to 100% occupancy in the active site. The Raman data also indicated that tazobactam forms a larger population of enamine than sulbactam or clavulanic acid does and that tazobactam's intermediate is also the most long-lived. The crystal structure provides a rationale for this finding since only tazobactam is able to form favorable intra- and intermolecular interactions in the active site that stabilize this trans-enamine intermediate. These interactions involve both the sulfone and triazolyl groups that distinguish tazobactam from clavulanic acid and sulbactam, respectively. The observed stabilization of the transient intermediate of tazobactam is thought to contribute to tazobactam's superior in vitro and in vivo clinical efficacy. Understanding the structural details of differing inhibitor effectiveness can aid the design of improved mechanism-based beta-lactamase inhibitors.
About this Structure
1RCJ is a Single protein structure of sequence from Klebsiella pneumoniae with and as ligands. Active as Beta-lactamase, with EC number 3.5.2.6 Full crystallographic information is available from OCA.
Reference
Tazobactam forms a stoichiometric trans-enamine intermediate in the E166A variant of SHV-1 beta-lactamase: 1.63 A crystal structure., Padayatti PS, Helfand MS, Totir MA, Carey MP, Hujer AM, Carey PR, Bonomo RA, van den Akker F, Biochemistry. 2004 Feb 3;43(4):843-8. PMID:14744126
Page seeded by OCA on Thu Feb 21 14:49:25 2008
Categories: Beta-lactamase | Klebsiella pneumoniae | Single protein | Akker, F van den. | Bonomo, R A. | Carey, M P. | Carey, P R. | Helfand, M S. | Hujer, A M. | Padayatti, P S. | Totir, M A. | MA4 | TBI | Beta-lactam hydrolase | Covalent intermediate | Detergent binding | Inhibitor design | Penicillinase
