Crosstalk between the protein surface and hydrophobic core in a core-swapped fibronectin type III domain |
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Authors: | Billings Kate S Best Robert B Rutherford Trevor J Clarke Jane |
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Affiliation: | 1 Cambridge University Chemical Laboratory, MRC Centre for Protein Engineering, Lensfield Road, Cambridge CB2 1EW, UK 2 Laboratory of Chemical Physics, NIDDK, National Institutes of Health, Building 5, Room 137A, Bethesda, MD 20892-0520, USA 3 MRC Centre for Protein Engineering, Hills Rd, Cambridge CB2 2QH, UK |
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Abstract: | Two homologous fibronectin type III (fnIII) domains, FNfn10 (the 10th fnIII domain of human fibronectin) and TNfn3 (the third fnIII domain of human tenascin), have essentially the same backbone structure, although they share only ∼ 24% sequence identity. While they share a similar folding mechanism with a common core of key residues in the folding transition state, they differ in many other physical properties. We use a chimeric protein, FNoTNc, to investigate the molecular basis for these differences. FNoTNc is a core-swapped protein, containing the “outside” (surface and loops) of FNfn10 and the hydrophobic core of TNfn3. Remarkably, FNoTNc retains the structure of the parent proteins despite the extent of redesign, allowing us to gain insight into which components of each parent protein are responsible for different aspects of its behaviour. Naively, one would expect properties that appear to depend principally on the core to be similar to TNfn3, for example, the response to mutations, folding kinetics and side-chain dynamics, while properties apparently determined by differences in the surface and loops, such as backbone dynamics, would be more like FNfn10. While this is broadly true, it is clear that there are also unexpected crosstalk effects between the core and the surface. For example, the anomalous response of FNfn10 to mutation is not solely a property of the core as we had previously suggested. |
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Keywords: | fnIII, fibronectin-type III FNfn10, 10th fnIII domain of human fibronectin TNfn3, third fnIII domain of human tenascin FNoTNc, a core-swapped protein with the &ldquo outside&rdquo (surface and loops) of FNfn10 and the core of TNfn3 GdmCl, guanidinium chloride HSQC, heteronuclear single quantum coherence TOCSY, total correlated spectroscopy |
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