Title
Long-range side-chain-main-chain interactions play crucial roles in stabilizing the (betaalpha)8 barrel motif of the alpha subunit of tryptophan synthase
UMMS Affiliation
Graduate School of Biomedical Sciences; Department of Biochemistry and Molecular Pharmacology
Publication Date
2007-06-26
Document Type
Article
Disciplines
Amino Acids, Peptides, and Proteins | Biochemistry | Genetic Phenomena | Inorganic Chemicals
Abstract
The role of hither-to-fore unrecognized long-range hydrogen bonds between main-chain amide hydrogens and polar side chains on the stability of a well-studied (betaalpha)8, TIM barrel protein, the alpha subunit of tryptophan synthase (alphaTS), was probed by mutational analysis. The F19-D46 and I97-D124 hydrogen bonds link the N terminus of a beta-strand with the C terminus of the succeeding antiparallel alpha-helix, and the A103-D130 hydrogen bond links the N terminus of an alpha-helix with the C terminus of the succeeding antiparallel beta-strand, forming clamps for the respective betaalpha or alphabeta hairpins. The individual replacement of these aspartic acid side chains with alanine leads to what appear to be closely related partially folded structures with significantly reduced far-UV CD ellipticity and thermodynamic stability. Comparisons with the effects of eliminating another main-chain-side-chain hydrogen bond, G26-S33, and two electrostatic side-chain-side-chain hydrogen bonds, D38-H92 and D112-H146, all in the same N-terminal folding unit of alphaTS, demonstrated a unique role for the clamp interactions in stabilizing the native barrel conformation. Because neither the asparagine nor glutamic acid variant at position 46 can completely reproduce the spectroscopic, thermodynamic, or kinetic folding properties of aspartic acid, both size and charge are crucial to its unique role in the clamp hydrogen bond. Kinetic studies suggest that the three clamp hydrogen bonds act in concert to stabilize the transition state leading to the fully folded TIM barrel motif.
Keywords
circular dichroism, protein folding, site-directed mutagenesis, thermodynamic and kinetic mechanisms, ba and ab hairpins
DOI of Published Version
10.1110/ps.062704507
Source
Protein Sci. 2007 Jul;16(7):1398-409. Link to article on publisher's site
Journal/Book/Conference Title
Protein science : a publication of the Protein Society
Related Resources
PubMed ID
17586773
Repository Citation
Yang X, Vadrevu R, Wu Y, Matthews CR. (2007). Long-range side-chain-main-chain interactions play crucial roles in stabilizing the (betaalpha)8 barrel motif of the alpha subunit of tryptophan synthase. Morningside Graduate School of Biomedical Sciences Student Publications. https://doi.org/10.1110/ps.062704507. Retrieved from https://escholarship.umassmed.edu/gsbs_sp/438