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Structural characterization of partially folded intermediates of apomyoglobin h64f

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Overview

authors

  • Schwarzinger, S.
  • Mohana-Borges, R.
  • Kroon, G. J. A.
  • Dyson, Jane
  • Wright, Peter

publication date

  • 2008

journal

  • Protein Science  Journal

abstract

  • We present a detailed investigation of unfolded and partially folded states of a mutant apomyoglobin (apoMb) where the distal histidine has been replaced by phenylalanine (H64F). Previous studies have shown that substitution of His64, located in the E helix of the native protein, stabilizes the equilibrium molten globule and native states and leads to an increase in folding rate and a change in the folding pathway. Analysis of changes in chemical shift and in backbone flexibility, detected via [1H]-15N heteronuclear nuclear Overhauser effect measurements, indicates that the phenylalanine substitution has only minor effects on the conformational ensemble in the acid- and urea-unfolded states, but has a substantial effect on the structure, dynamics, and stability of the equilibrium molten globule intermediate formed near pH 4. In H64F apomyoglobin, additional regions of the polypeptide chain are recruited into the compact core of the molten globule. Since the phenylalanine substitution has negligible effect on the unfolded ensemble, its influence on folding rate and stability comes entirely from interactions within the compact folded or partly folded states. Replacement of His64 with Phe leads to favorable hydrophobic packing between the helix E region and the molten globule core and leads to stabilization of helix E secondary structure and overall thermodynamic stabilization of the molten globule. The secondary structure of the equilibrium molten globule parallels that of the burst phase kinetic intermediate; both intermediates contain significant helical structure in regions of the polypeptide that comprise the A, B, E, G, and H helices of the fully folded protein.

subject areas

  • Amino Acid Substitution
  • Apoproteins
  • Hydrogen-Ion Concentration
  • Mutant Proteins
  • Myoglobin
  • Protein Conformation
  • Protein Folding
  • Protein Structure, Secondary
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Research

keywords

  • NMR spectroscopy
  • chain dynamics
  • hydrophobic interaction
  • molten globule
  • myoglobin
  • protein folding
  • protein stability
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Identity

PubMed Central ID

  • PMC2222724

International Standard Serial Number (ISSN)

  • 0961-8368

Digital Object Identifier (DOI)

  • 10.1110/ps.073187208

PubMed ID

  • 18227434
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Additional Document Info

start page

  • 313

end page

  • 321

volume

  • 17

issue

  • 2

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