Stalled flavodoxin binds its cofactor while fully exposed outside the ribosome.
| Autor: | Houwman JA; Laboratory of Biochemistry, Wageningen University, Dreijenlaan 3, 6703 HA Wageningen, the Netherlands., Westphal AH; Laboratory of Biochemistry, Wageningen University, Dreijenlaan 3, 6703 HA Wageningen, the Netherlands., van Berkel WJ; Laboratory of Biochemistry, Wageningen University, Dreijenlaan 3, 6703 HA Wageningen, the Netherlands., van Mierlo CP; Laboratory of Biochemistry, Wageningen University, Dreijenlaan 3, 6703 HA Wageningen, the Netherlands. Electronic address: carlo.vanmierlo@wur.nl. |
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| Jazyk: | angličtina |
| Zdroj: | Biochimica et biophysica acta [Biochim Biophys Acta] 2015 Oct; Vol. 1854 (10 Pt A), pp. 1317-24. Date of Electronic Publication: 2015 Jun 11. |
| DOI: | 10.1016/j.bbapap.2015.06.004 |
| Abstrakt: | Correct folding of proteins is crucial for cellular homeostasis. More than thirty percent of proteins contain one or more cofactors, but the impact of these cofactors on co-translational folding remains largely unknown. Here, we address the binding of flavin mononucleotide (FMN) to nascent flavodoxin, by generating ribosome-arrested nascent chains that expose either the entire protein or C-terminally truncated segments thereof. The native α/β parallel fold of flavodoxin is among the most ancestral and widely distributed folds in nature and exploring its co-translational folding is thus highly relevant. In Escherichia coli (strain BL21(DE3) Δtig::kan) FMN turns out to be limiting for saturation of this flavoprotein on time-scales vastly exceeding those of flavodoxin synthesis. Because the ribosome affects protein folding, apoflavodoxin cannot bind FMN during its translation. As a result, binding of cofactor to released protein is the last step in production of this flavoprotein in the cell. We show that once apoflavodoxin is entirely synthesized and exposed outside the ribosome to which it is stalled by an artificial linker containing the SecM sequence, the protein is natively folded and capable of binding FMN. (Copyright © 2015. Published by Elsevier B.V.) |
| Databáze: | MEDLINE |
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