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Promiscuous contacts and heightened dynamics increase thermostability in an engineered variant of the engrailed
Michelle E McCully1, David A C Beck, Valerie Daggett
1Biomolecular Structure and Design Program, University of Washington, Seattle, WA 98195-5013, USA.
Abstract:
A thermostabilized variant (UVF) of the engrailed homeodomain (EnHD) was previously engineered by Mayo and co-workers. The melting temperature of the non-natural, designed protein is 50°C higher than the natural wild-type protein (>99 vs. 52°C), and the two proteins share 22% sequence identity. We have performed extensive (1 μs) all-atom, explicit solvent molecular dynamics simulations of the wild-type and engineered proteins to investigate their structural and dynamic properties at room temperature and at 100°C. Our simulations are in good agreement with nuclear magnetic resonance data available for the two proteins [nuclear Overhauser effect crosspeaks (NOEs), J-coupling constants and order parameters for EnHD; and NOEs for UVF], showing that we reproduce the backbone dynamics and side chain packing in the native state of both proteins. UVF was more dynamic at room temperature than EnHD, with respect to both its backbone and side chain motion. When the temperature was raised, the thermostable protein maintained this mobility while retaining its native conformation. EnHD, on the other hand, was unable to maintain its more rigid native structure at higher temperature and began to unfold. Heightened protein dynamics leading to promiscuous and dynamically interchangeable amino acid contacts makes UVF more tolerant to increasing temperature, providing a molecular explanation for heightened thermostability of this protein.
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