title: Urea-induced unfolding of the alpha subunit of tryptophan synthase: one-dimensional proton NMR evidence for residual structure near histidine-92 at high denaturant concentration.
abstract: The urea-induced unfolding reaction of the alpha subunit of tryptophan synthase was monitored by examining the chemical shifts and peak areas of the C epsilon protons of the four histidine residues with 1D NMR spectroscopy. In a native base-line region defined by tyrosine absorbance and far-UV circular dichroism spectroscopy, histidine-146 appears to undergo a rapid, local unfolding reaction at increasing denaturant concentrations. As the native form is converted to a previously detected stable intermediate between 2 and 3 M urea [Matthews, C. R., & Crisanti, M. M. (1981) Biochemistry 20, 784], histidines-92 and -146 in the amino folding unit (residues 1-188) and histidines-195 and -244 in the carboxy folding unit (residues 189-268) all experience a change in their environments which is slow on the NMR time scale. The subsequent conversion of this intermediate to a newly detected, stable, partially folded form populated at 5 M urea appears to have no effect on any of the histidines at 25 degrees C when an intermolecular association process involving His-244 is taken into account. Strikingly, a slow exchange process involving only His-92 is observed to begin at 5 M urea where the unfolding transitions monitored by absorbance or far-UV circular dichroism spectroscopy are essentially complete. This residual tertiary structure unfolds in a cooperative fashion as the urea concentration is increased to 8 M.
authors.affiliation: Department of Chemistry, Pennsylvania State University, University Park 16802.
authors.forename: G
authors.initials: G
authors.lastname: Saab-Rincón
authors.forename: C L
authors.initials: CL
authors.lastname: Froebe
authors.forename: C R
authors.initials: CR
authors.lastname: Matthews
mesh_terms: D000595:Amino Acid Sequence; D004926:Escherichia coli / Q000201:enzymology; D006639:Histidine / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry*; D011489:Protein Denaturation; D017510:Protein Folding; D011522:Protons; D013816:Thermodynamics; D014367:Tryptophan Synthase / Q000737:chemistry*; D014508:Urea / Q000737:chemistry*
chemical_list: D010446:Peptide Fragments; D011522:Protons; D006639:Histidine; D014508:Urea; D014367:Tryptophan Synthase