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Modification of residue 42 of the active site loop with a lysinemimetic sidechain rescues isochorismate-pyruvate lyase activity in Pseudomonas aeruginosa PchB
dc.contributor.author | Olucha, José | |
dc.contributor.author | Meneely, Kathleen M. | |
dc.contributor.author | Lamb, Audrey L. | |
dc.date.accessioned | 2017-04-14T19:27:06Z | |
dc.date.available | 2017-04-14T19:27:06Z | |
dc.date.issued | 2012-09-25 | |
dc.identifier.citation | Olucha, J., Meneely, K. M., & Lamb, A. L. (2012). Modification of residue 42 of the active site loop with a lysine-mimetic sidechain rescues isochorismate-pyruvate lyase activity in Pseudomonas aeruginosa PchB. Biochemistry, 51(38), 7525–7532. http://doi.org/10.1021/bi300472n | en_US |
dc.identifier.uri | http://hdl.handle.net/1808/23700 | |
dc.description.abstract | PchB is an isochorismate-pyruvate lyase from Pseudomonas aeruginosa. A positively charged lysine residue is located in a flexible loop that behaves as a lid to the active site, and the lysine residue is required for efficient production of salicylate. A variant of PchB that lacks the lysine at residue 42 has a reduced catalytic free energy of activation of up to 4.4 kcal/mol. Construction of a lysine isosteric residue bearing a positive charge at the appropriate position leads to the recovery of 2.5–2.7 kcal/mol (about 60%) of the 4.4 kcal/mol by chemical rescue. Exogenous addition of ethylamine to the K42A variant leads to a neglible recovery of activity (0.180 kcal/mol, roughly 7% rescue), whereas addition of propylamine caused an additional modest loss in catalytic power (0.056 kcal/mol, or −2% loss). This is consistent with the view that (a) the lysine-42 residue is required in a specific conformation to stabilize the transition state and (b) the correct conformation is achieved for a lysine-mimetic sidechain at site 42 in the course of loop closure, as expected for transition-state stabilization by the side chain ammonio-function. That the positive charge is the main effector of transition state stabilization is shown by the construction of a lysine-isosteric residue capable of exerting steric effects and hydrogen bonding but not electrostatic effects, leading to a modest increase of catalytic power (0.267 – 0.505 kcal/mol of catalytic free energy, or roughly 6 – 11% rescue). | en_US |
dc.publisher | ACS | en_US |
dc.rights | Copyright © 2012 American Chemical Society | en_US |
dc.title | Modification of residue 42 of the active site loop with a lysinemimetic sidechain rescues isochorismate-pyruvate lyase activity in Pseudomonas aeruginosa PchB | en_US |
dc.type | Article | en_US |
kusw.kuauthor | Meneely, Kathleen M. | |
kusw.kuauthor | Lamb, Audrey L. | |
kusw.kudepartment | Higuchi Biosciences Center | en_US |
kusw.kudepartment | Molecular Biosciences | en_US |
dc.identifier.doi | 10.1021/bi300472n | en_US |
kusw.oaversion | Scholarly/refereed, author accepted manuscript | en_US |
kusw.oapolicy | This item meets KU Open Access policy criteria. | en_US |
dc.rights.accessrights | openAccess |