title: Dynamics of compact denatured states of glutaminyl-tRNA synthetase probed by bis-ANS binding kinetics.
abstract: Bis-ANS binds to native glutaminyl-tRNA synthetase (GlnRS) with a fast and a slow phase. The rate constant of the slow phase is independent of bis-ANS concentration suggesting a slow conformational change in the pathway of bis-ANS binding. Aging of GlnRS causes a large decrease of the slow phase amplitude with concomitant increase of the fast phase amplitude. Several other large, multi-domain proteins show similar patterns upon aging. The near UV-CD spectra of the native and the aged GlnRS remain similar. Significant changes in far UV-CD, acrylamide quenching and sulfhydryl reactivity, are seen upon aging, suggesting disruptions in native interactions. Refolding of GlnRS from the urea-denatured state rapidly produces a state that is very similar to the equilibrium molten globule state. Bis-ANS binds to the molten globule state with kinetics similar to that of the aged state and unlike that of the native state. This suggests that the slow binding phase of bis-ANS, seen in native proteins, originate from relatively high energy barriers between the native and the more open states. Thus bis-ANS can be used as a powerful probe for large amplitude, low-frequency motions of proteins.
authors: Department of Biophysics, Bose Institute, Calcutta, India.
authors: A
authors: A
authors: Bhattacharyya
authors: A K
authors: AK
authors: Mandal
authors: R
authors: R
authors: Banerjee
authors: S
authors: S
authors: Roy
mesh_terms: D000604:Amino Acyl-tRNA Synthetases / Q000737:chemistry* / Q000378:metabolism*; D000817:Anilino Naphthalenesulfonates; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002942:Circular Dichroism; D005456:Fluorescent Dyes; D066298:In Vitro Techniques; D007700:Kinetics; D011487:Protein Conformation; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D013816:Thermodynamics
chemical_list: D000817:Anilino Naphthalenesulfonates; D005456:Fluorescent Dyes; C035763:5,5'-bis(8-(phenylamino)-1-naphthalenesulfonate); D000604:Amino Acyl-tRNA Synthetases; C020659:glutaminyl-tRNA synthetase