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Probing the non-native H helix translocation in apomyoglobin folding intermediates.
Biochemistry. 2014 Jun 17; 53(23):3767-80.B

Abstract

Apomyoglobin folds via sequential helical intermediates that are formed by rapid collapse of the A, B, G, and H helix regions. An equilibrium molten globule with a similar structure is formed near pH 4. Previous studies suggested that the folding intermediates are kinetically trapped states in which folding is impeded by non-native packing of the G and H helices. Fluorescence spectra of mutant proteins in which cysteine residues were introduced at several positions in the G and H helices show differential quenching of W14 fluorescence, providing direct evidence of translocation of the H helix relative to helices A and G in both the kinetic and equilibrium intermediates. Förster resonance energy transfer measurements show that a 5-({2-[(acetyl)amino]ethyl}amino)naphthalene-1-sulfonic acid acceptor coupled to K140C (helix H) is closer to Trp14 (helix A) in the equilibrium molten globule than in the native state, by a distance that is consistent with sliding of the H helix in an N-terminal direction by approximately one helical turn. Formation of an S108C-L135C disulfide prevents H helix translocation in the equilibrium molten globule by locking the G and H helices into their native register. By enforcing nativelike packing of the A, G, and H helices, the disulfide resolves local energetic frustration and facilitates transient docking of the E helix region onto the hydrophobic core but has only a small effect on the refolding rate. The apomyoglobin folding landscape is highly rugged, with several energetic bottlenecks that frustrate folding; relief of any one of the major identified bottlenecks is insufficient to speed progression to the transition state.

Authors+Show Affiliations

Department of Molecular Biology and Skaggs Institute of Chemical Biology, The Scripps Research Institute , 10550 North Torrey Pines Road, La Jolla, California 92037, United States.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural

Language

eng

PubMed ID

24857522

Citation

Aoto, Phillip C., et al. "Probing the Non-native H Helix Translocation in Apomyoglobin Folding Intermediates." Biochemistry, vol. 53, no. 23, 2014, pp. 3767-80.
Aoto PC, Nishimura C, Dyson HJ, et al. Probing the non-native H helix translocation in apomyoglobin folding intermediates. Biochemistry. 2014;53(23):3767-80.
Aoto, P. C., Nishimura, C., Dyson, H. J., & Wright, P. E. (2014). Probing the non-native H helix translocation in apomyoglobin folding intermediates. Biochemistry, 53(23), 3767-80. https://doi.org/10.1021/bi500478m
Aoto PC, et al. Probing the Non-native H Helix Translocation in Apomyoglobin Folding Intermediates. Biochemistry. 2014 Jun 17;53(23):3767-80. PubMed PMID: 24857522.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Probing the non-native H helix translocation in apomyoglobin folding intermediates. AU - Aoto,Phillip C, AU - Nishimura,Chiaki, AU - Dyson,H Jane, AU - Wright,Peter E, Y1 - 2014/06/04/ PY - 2014/5/27/entrez PY - 2014/5/27/pubmed PY - 2014/9/3/medline SP - 3767 EP - 80 JF - Biochemistry JO - Biochemistry VL - 53 IS - 23 N2 - Apomyoglobin folds via sequential helical intermediates that are formed by rapid collapse of the A, B, G, and H helix regions. An equilibrium molten globule with a similar structure is formed near pH 4. Previous studies suggested that the folding intermediates are kinetically trapped states in which folding is impeded by non-native packing of the G and H helices. Fluorescence spectra of mutant proteins in which cysteine residues were introduced at several positions in the G and H helices show differential quenching of W14 fluorescence, providing direct evidence of translocation of the H helix relative to helices A and G in both the kinetic and equilibrium intermediates. Förster resonance energy transfer measurements show that a 5-({2-[(acetyl)amino]ethyl}amino)naphthalene-1-sulfonic acid acceptor coupled to K140C (helix H) is closer to Trp14 (helix A) in the equilibrium molten globule than in the native state, by a distance that is consistent with sliding of the H helix in an N-terminal direction by approximately one helical turn. Formation of an S108C-L135C disulfide prevents H helix translocation in the equilibrium molten globule by locking the G and H helices into their native register. By enforcing nativelike packing of the A, G, and H helices, the disulfide resolves local energetic frustration and facilitates transient docking of the E helix region onto the hydrophobic core but has only a small effect on the refolding rate. The apomyoglobin folding landscape is highly rugged, with several energetic bottlenecks that frustrate folding; relief of any one of the major identified bottlenecks is insufficient to speed progression to the transition state. SN - 1520-4995 UR - https://www.unboundmedicine.com/medline/citation/24857522/Probing_the_non_native_H_helix_translocation_in_apomyoglobin_folding_intermediates_ L2 - https://doi.org/10.1021/bi500478m DB - PRIME DP - Unbound Medicine ER -