title: NMR and CD spectroscopy show that imino acid restriction of the unfolded state leads to efficient folding.
abstract: Protein folding is determined by molecular features in the unfolded state, as well as the native folded structure. In the unfolded state, imino acids both restrict conformational space and present cis-trans isomerization barriers to folding. Because of its high proline and hydroxyproline content, the collagen triple-helix offers an opportunity to characterize the impact of imino acids on the unfolded state and folding kinetics. Here, NMR and CD spectroscopy are used to characterize the role of imino acids in a triple-helical peptide, T1-892, which contains an 18-residue sequence from type I collagen and a C-terminal (Gly-Pro-Hyp)(4) domain. The replacement of Pro or Hyp by an Ala in the (Gly-Pro-Hyp)(4) region significantly decreases the folding rate at low but not high concentrations, consistent with less efficient nucleation. To understand the molecular basis of the decreased folding rate, changes in the unfolded as well as the folded states of the peptides were characterized. While the trimer states of the peptides are all similar, NMR dynamics studies show monomers with all trans (Gly-Pro-Hyp)(4) are less flexible than monomers containing Pro --> Ala or Hyp --> Ala substitutions. Nucleation requires all trans bonds in the (Gly-Pro-Hyp)(4) domain and the constrained monomer state of the all trans nucleation domain in T1-892 increases its competency to initiate triple-helix formation and illustrates the impact of the unfolded state on folding kinetics.
authors.affiliation: Department of Biochemistry, Robert-Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Piscataway, New Jersey 08854, USA.
authors.forename: Yujia
authors.initials: Y
authors.lastname: Xu
authors.forename: Timothy
authors.initials: T
authors.lastname: Hyde
authors.forename: Xin
authors.initials: X
authors.lastname: Wang
authors.forename: Manjiri
authors.initials: M
authors.lastname: Bhate
authors.forename: Barbara
authors.initials: B
authors.lastname: Brodsky
authors.forename: Jean
authors.initials: J
authors.lastname: Baum
mesh_terms: D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D002942:Circular Dichroism / Q000379:methods*; D024042:Collagen Type I / Q000737:chemistry; D019281:Dimerization; D006909:Hydroxyproline / Q000737:chemistry; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008956:Models, Chemical; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011392:Proline / Q000737:chemistry; D011487:Protein Conformation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013053:Spectrophotometry; D013696:Temperature; D013816:Thermodynamics; D013997:Time Factors
chemical_list: D000596:Amino Acids; D024042:Collagen Type I; D010455:Peptides; D011392:Proline; D000409:Alanine; D006909:Hydroxyproline