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The determinants of stability and folding in evolutionarily diverged cytochromes c.

ABSTRACT: Cytochrome c has served as a paradigm for the study of protein stability, folding, and molecular evolution, but it remains unclear how these aspects of the protein are related. For example, while the bovine and equine cytochromes c are known to have different stabilities, and possibly different folding mechanisms, it is not known how these differences arise from just three amino acid substitutions introduced during divergence. Using site-selectively incorporated carbon-deuterium bonds, we show that like the equine protein, bovine cytochrome c is induced to unfold by guanidine hydrochloride via a stepwise mechanism, but it does not populate an intermediate as is observed with the equine protein. The increased stability also results in more similar free energies of unfolding observed at different sites within the protein, giving the appearance of a more concerted mechanism. Furthermore, we show that the differences in stability and folding appear to result from a single amino acid substitution that stabilizes a helix by allowing for increased solvation of its N-terminus.

SUBMITTER: Thielges MC

PROVIDER: S-EPMC2990880 | biostudies-literature | 2009 Apr

REPOSITORIES: biostudies-literature

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The determinants of stability and folding in evolutionarily diverged cytochromes c.

Thielges Megan C MC Zimmermann Jörg J Dawson Philip E PE Romesberg Floyd E FE

Journal of molecular biology 20090304 1

Cytochrome c has served as a paradigm for the study of protein stability, folding, and molecular evolution, but it remains unclear how these aspects of the protein are related. For example, while the bovine and equine cytochromes c are known to have different stabilities, and possibly different folding mechanisms, it is not known how these differences arise from just three amino acid substitutions introduced during divergence. Using site-selectively incorporated carbon-deuterium bonds, we show t ...[more]

PMID: 19268474

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Project description:BACKGROUND: Gcn5 is a transcriptional coactivator with histone acetyltransferase activity that is conserved with regard to structure as well as its histone substrates throughout the eukaryotes. Gene regulatory networks within cells are thought to be evolutionarily diverged. The use of evolutionarily divergent yeast species, such as S. cerevisiae and S. pombe, which can be studied under similar environmental conditions, provides an opportunity to examine the interface between conserved regulatory components and their cellular applications in different organisms. RESULTS: We show that Gcn5 is important for a common set of stress responses in evolutionarily diverged yeast species and that the activity of the conserved histone acetyltransferase domain is required. We define a group of KCl stress response genes in S. cerevisiae that are specifically dependent on Gcn5. Gcn5 is localised to many Gcn5-dependent genes including Gcn5 repressed targets such as FLO8. Gcn5 regulates divergent sets of KCl responsive genes in S. cerevisiae and S. pombe. Genome-wide localization studies showed a tendency for redistribution of Gcn5 during KCl stress adaptation in S. cerevisiae from short genes to the transcribed regions of long genes. An analogous redistribution was not observed in S. pombe. CONCLUSIONS: Gcn5 is required for the regulation of divergent sets of KCl stress-response genes in S. cerevisiae and S. pombe even though it is required a common group of stress responses, including the response to KCl. Genes that are physically associated with Gcn5 require its activity for their repression or activation during stress adaptation, providing support for a role of Gcn5 as a corepressor as well as a coactivator. The tendency of Gcn5 to re-localise to the transcribed regions of long genes during KCl stress adaptation suggests that Gcn5 plays a specific role in the expression of long genes under adaptive conditions, perhaps by regulating transcriptional elongation as has been seen for Gcn5 in S. pombe. Interestingly an analogous redistribution of Gcn5 is not seen in S. pombe. The study thus provides important new insights in relation to why coregulators like Gcn5 are required for the correct expression of some genes but not others.

Dataset Information

The determinants of stability and folding in evolutionarily diverged cytochromes c.

Publications

The determinants of stability and folding in evolutionarily diverged cytochromes c.

Similar Datasets

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