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Cavagnero, Silvia ; Nishimura, Chiaki ; Schwarzinger, Stephan ; Dyson, H. Jane ; Wright, Peter E.:
Conformational and Dynamical Characterization of the Molten Globule State of an Apomyoglobin Mutant with an Altered Folding Pathway.
In: Biochemistry.
Bd. 40
(2001)
Heft 48
.
- S. 14459-14467.
ISSN 1520-4995
DOI: https://doi.org/10.1021/bi011500n
Abstract
Kinetic and equilibrium studies of apomyoglobin folding pathways and intermediates have provided important insights into the mechanism of protein folding. To investigate the role of intrinsic helical propensities in the apomyoglobin folding process, a mutant has been prepared in which Asn132 and Glu136 have been substituted with glycine to destabilize the H helix. The structure and dynamics of the equilibrium molten globule state formed at pH 4.1 have been examined using NMR spectroscopy. Deviations of backbone (13)C(alpha) and (13)CO chemical shifts from random coil values reveal high populations of helical structure in the A and G helix regions and in part of the B helix. However, the H helix is significantly destabilized compared to the wild-type molten globule. Heteronuclear [(1)H]-(15)N NOEs show that, although the polypeptide backbone in the H helix region is more flexible than in the wild-type protein, its motions are restricted by transient hydrophobic interactions with the molten globule core. Quench flow hydrogen exchange measurements reveal stable helical structure in the A and G helices and part of the B helix in the burst phase kinetic intermediate and confirm that the H helix is largely unstructured. Stabilization of structure in the H helix occurs during the slow folding phases, in synchrony with the C and E helices and the CD region. The kinetic and equilibrium molten globule intermediates formed by N132G/E136G are similar in structure. Although both the wild-type apomyoglobin and the mutant fold via compact helical intermediates, the structures of the intermediates and consequently the detailed folding pathways differ. Apomyoglobin is therefore capable of compensating for mutations by using alternative folding pathways within a common basic framework. Tertiary hydrophobic interactions appear to play an important role in the formation and stabilization of secondary structure in the H helix of the N132G/E136G mutant. These studies provide important insights into the interplay between secondary and tertiary structure formation in protein folding.
Weitere Angaben
| Publikationsform: | Artikel in einer Zeitschrift |
|---|---|
| Begutachteter Beitrag: | Ja |
| Institutionen der Universität: | Fakultäten > Fakultät für Biologie, Chemie und Geowissenschaften Fakultäten > Fakultät für Biologie, Chemie und Geowissenschaften > Fachgruppe Chemie Fakultäten |
| Titel an der UBT entstanden: | Ja |
| Themengebiete aus DDC: | 500 Naturwissenschaften und Mathematik > 500 Naturwissenschaften |
| Eingestellt am: | 06 Aug 2019 06:45 |
| Letzte Änderung: | 13 Jul 2021 13:23 |
| URI: | https://eref.uni-bayreuth.de/id/eprint/51779 |