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Dissection of a (βα)8-barrel enzyme into two folded halves

Nature Structural Biology volume 8, pages 32–36 (2001)Cite this article

Abstract

The (βα)8-barrel, which is the most frequently encountered protein fold, is generally considered to consist of a single structural domain. However, the X-ray structure of the imidazoleglycerol phosphate synthase (HisF) from Thermotoga maritima has identified it as a (βα)8-barrel made up of two superimposable subdomains (HisF-N and HisF-C). HisF-N consists of the four N-terminal (βα) units and HisF-C of the four C-terminal (βα) units. It has been postulated, therefore, that HisF evolved by tandem duplication and fusion from an ancestral half-barrel. To test this hypothesis, HisF-N and HisF-C were produced in Escherichia coli, purified and characterized. Separately, HisF-N and HisF-C are folded proteins, but are catalytically inactive. Upon co-expression in vivo or joint refolding in vitro, HisF-N and HisF-C assemble to the stoichiometric and catalytically fully active HisF-NC complex. These findings support the hypothesis that the (βα)8-barrel of HisF evolved from an ancestral half-barrel and have implications for the folding mechanism of the members of this large protein family.

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Figure 1: The N-terminal and C-terminal halves of HisF have very similar structures.
Figure 2: CD spectra of HisF-Nw, HisF-C and HisF-NC are indicative of native secondary and tertiary structures.
Figure 3: HisF-C is more resistant to proteolysis than HisF-N, and complex formation slows down the degradation of both fragments.
Figure 4: Models of the quaternary structures of the predominant association states of HisF, and of HisF-N, HisF-C and HisF-NC.

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Acknowledgements

We thank H.-J. Fritz, K. Kirschner, D. Lang, R. Thoma and M. Wilmanns for support and helpful discussions. This work was sponsored by grants from the Deutsche Forschungsgemeinschaft and a Heisenberg-fellowship to R.S. This paper is dedicated to Professor Marianne Baudler on the occasion of her 80th birthday.

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Authors and Affiliations

  1. Abteilung Molekulare Genetik und Präparative Molekularbiologie, Institut für Mikrobiologie und Genetik, Georg-August-Universität Göttingen, Grisebachstr. 8, Göttingen, D-37077, Germany

    Birte Höcker, Silke Beismann-Driemeyer, Stefan Hettwer & Reinhard Sterner

  2. Universität zu Köln, Institut für Biochemie , Otto-Fischer-Str. 12-14, Köln, D-50674, Germany

    Birte Höcker, Silke Beismann-Driemeyer, Stefan Hettwer & Reinhard Sterner

  3. Abteilung für Biophysikalische Chemie, Biozentrum der Universität Basel, Klingelbergstr. 70, Basel, CH-4056, Switzerland

    Ariel Lustig

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Correspondence to Reinhard Sterner.

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Höcker, B., Beismann-Driemeyer, S., Hettwer, S. et al. Dissection of a (βα)8-barrel enzyme into two folded halves. Nat Struct Mol Biol 8, 32–36 (2001). https://doi.org/10.1038/83021

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