1a5l: Difference between revisions
No edit summary |
No edit summary |
||
| Line 1: | Line 1: | ||
[[Image:1a5l.gif|left|200px]] | [[Image:1a5l.gif|left|200px]] | ||
'''K217C VARIANT OF KLEBSIELLA AEROGENES UREASE''' | {{Structure | ||
|PDB= 1a5l |SIZE=350|CAPTION= <scene name='initialview01'>1a5l</scene>, resolution 2.2Å | |||
|SITE= | |||
|LIGAND= | |||
|ACTIVITY= [http://en.wikipedia.org/wiki/Urease Urease], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.5.1.5 3.5.1.5] | |||
|GENE= UREA, UREB, UREC ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=28451 Klebsiella aerogenes]) | |||
}} | |||
'''K217C VARIANT OF KLEBSIELLA AEROGENES UREASE''' | |||
==Overview== | ==Overview== | ||
| Line 7: | Line 16: | ||
==About this Structure== | ==About this Structure== | ||
1A5L is a [ | 1A5L is a [[Protein complex]] structure of sequences from [http://en.wikipedia.org/wiki/Klebsiella_aerogenes Klebsiella aerogenes]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1A5L OCA]. | ||
==Reference== | ==Reference== | ||
Chemical rescue of Klebsiella aerogenes urease variants lacking the carbamylated-lysine nickel ligand., Pearson MA, Schaller RA, Michel LO, Karplus PA, Hausinger RP, Biochemistry. 1998 Apr 28;37(17):6214-20. PMID:[http:// | Chemical rescue of Klebsiella aerogenes urease variants lacking the carbamylated-lysine nickel ligand., Pearson MA, Schaller RA, Michel LO, Karplus PA, Hausinger RP, Biochemistry. 1998 Apr 28;37(17):6214-20. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/9558361 9558361] | ||
[[Category: Klebsiella aerogenes]] | [[Category: Klebsiella aerogenes]] | ||
[[Category: Protein complex]] | [[Category: Protein complex]] | ||
| Line 23: | Line 32: | ||
[[Category: nickel metalloenzyme]] | [[Category: nickel metalloenzyme]] | ||
''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Mar 20 09:53:15 2008'' | ||
Revision as of 10:53, 20 March 2008
| |||||||
| , resolution 2.2Å | |||||||
|---|---|---|---|---|---|---|---|
| Gene: | UREA, UREB, UREC (Klebsiella aerogenes) | ||||||
| Activity: | Urease, with EC number 3.5.1.5 | ||||||
| Coordinates: | save as pdb, mmCIF, xml | ||||||
K217C VARIANT OF KLEBSIELLA AEROGENES UREASE
OverviewOverview
Klebsiella aerogenes urease possesses a dinuclear metallocenter in which two nickel atoms are bridged by carbamylated Lys217. To assess whether carbamate-specific chemistry is required for urease activity, site-directed mutagenesis and chemical rescue strategies were combined in efforts to place a carboxylate group at the location of this metal ligand. Urease variants with Lys217 replaced by Glu, Cys, and Ala (K217E, K217C/C319A, and K217A proteins) were purified, shown to be activated by incubation with small organic acids plus Ni(II), and structurally characterized. K217C/C319A urease possessed a second change in which Cys319 was replaced by Ala in order to facilitate efforts to chemically modify Cys217; however, this covalent modification approach did not produce active urease. Chemical rescue of the K217E, K217C/C319A, and K217A variants required 2, 2, and 10 h, respectively, to reach maximal activity levels. The highest activity generated [224 micromol of urea degraded.min-1.(mg of protein)-1, for K217C/C319A urease incubated with 500 mM formic acid and 10 mM Ni at pH 6.5] corresponded to 56% of that measured for in vitro activation of the wild-type apoprotein. While the K217E apoprotein showed minimal structural perturbations, the K217C/C319A apoprotein showed a disordering of some active site residues, and the K217A apoprotein revealed a repositioning of His219 to allow the formation of a hydrogen bond with Thr169, thus replacing the hydrogen bond between the amino group of Lys217 and Thr169 in the native enzyme. Importantly, these structures allow rationalization of the relative rates and yields of chemical rescue experiments. The crystal structures of chemically rescued K217A and K217C/C319A ureases revealed a return of the active site residues to their wild-type positions. In both cases, noncovalently bound formate was structurally equivalent to the Lys-carbamate as the bridging metallocenter ligand. We conclude that carbamate-specific chemistry is not required for urease catalysis.
About this StructureAbout this Structure
1A5L is a Protein complex structure of sequences from Klebsiella aerogenes. Full crystallographic information is available from OCA.
ReferenceReference
Chemical rescue of Klebsiella aerogenes urease variants lacking the carbamylated-lysine nickel ligand., Pearson MA, Schaller RA, Michel LO, Karplus PA, Hausinger RP, Biochemistry. 1998 Apr 28;37(17):6214-20. PMID:9558361
Page seeded by OCA on Thu Mar 20 09:53:15 2008