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UNIVERSITY OF BUCHAREST
FACULTY OF PHYSICS Guest
2024-11-22 1:39 |
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Conference: Bucharest University Faculty of Physics 2005 Meeting
Section: Electricity and Biophysics
Title:
Fluorescent Study of the Interaction between
Strepococcocus aureus UMP Kinase with UTP !
Authors:
Claudia Firanescu1, Doina Gazdaru1, Nadia Bucurenci2 and Aurel Popescu1
Affiliation:
1Department of Electricity and Biophysics, Faculty of Physics, University of Bucharest
2Institute “Ion Cantacuzino”, Bucharest of Microbioplogy
E-mail
claudiafir@yahoo.com
Keywords:
Strepococcocus aureus UMP Kinase, UTP, fluorescence
Abstract:
Fluorescent techniques can provide valuable physical parameters consistent with those obtained by the crystallographic methods. The various environments of the fluorophores of a folded protein and the unique stereochemistry of the protein chain affect the fluorophore state in many ways, allowing us to characterize and follow the changes in the protein folding. Fluorescence-based techniques applied to the proteins are giving information on their binding sites, interactions with solvent, the degrees of flexibility, internal motions, rotational diffusion coefficients, etc. We tested the fluorescent behavior of one member of NMP kinase family, namely, Streptococcus aureus UMP kinase. NMP kinases are ubiquitous enzymes of the living cells, very important in the cellular energetic metabolism, and also in synthesis of nucleic acid precursors. Is seems that Streptococcus aureus UMP kinase has a different behavior, as compared to those of UMP kinase from Streptococcus pneumoniae or from Escherichia coli. The binding of a specific ligand, UTP, to this UMP kinase is seems to be a cooperative process, being accompanied by an 8.2-fold decrease in the intrinsic fluorescence, on a UTP concentration range between 50 and 1500 mM. UMP kinase from Streptococcus aureus is exhibiting an intrinsic fluorescence, due to its tryptophan, that is decreasing with the increase of UTP concentration. By the ipothesys of a collisional quenching of fluorescence, the fluorescence data were fitted by Stern-Volmer equation. It was obtained a very large Stern-Volmer constant of 5 x 103 M-1. Using a mean lifetime fluorescence of tryptophan, in neutral aqueous solutions, we found a biomolecular quenching constant, kq, of 2 × 1012 M-1 sec-1. This value is larger than that possible for a diffusion-controlled reaction, and therefore complex formation is suspected.
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