Mapping the Conformational Stability of Maltose Binding Protein at the Residue Scale Using Nuclear Magnetic Resonance Hydrogen Exchange Experiments

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• 作者：C茅line Merstorf ; Olek Maciejak ; J茅r么me Math茅 ; Manuela Pastoriza-Gallego ; B茅n茅dicte Thiebot ; Marie-Jeanne Cl茅ment ; Juan Pelta ; Lo茂c Auvray ; Patrick A. Curmi ; Philippe Savarin
• 刊名：Biochemistry
• 出版年：2012
• 出版时间：November 6, 2012
• 年：2012
• 卷：51
• 期：44
• 页码：8919-8930
• 全文大小：614K
• 年卷期：v.51,no.44(November 6, 2012)
• ISSN：1520-4995

文摘

Being able to differentiate local fluctuations from global folding鈥搖nfolding dynamics of a protein is of major interest for improving our understanding of structure鈥揻unction determinants. The maltose binding protein (MBP), a protein that belongs to the maltose transport system, has a structure composed of two globular domains separated by a rigid-body 鈥渉inge bending鈥? Here we determined, by using hydrogen exchange (HX) nuclear magnetic resonance experiments, the apparent stabilization free energies of 101 residues of MBP bound to 尾-cyclodextrin (MBP鈭捨睠D) under native conditions. We observed that the last helix of MBP (helix 伪14) has a lower protection factor than the rest of the protein. Further, HX experiments were performed using guanidine hydrochloride under subdenaturing conditions to discriminate between local fluctuations and global unfolding events and to determine the MBP鈭捨睠D energy landscape. The results show that helix 伪4 and a part of helices 伪5 and 伪6 are clearly grouped into a subdenaturing folding unit and represent a partially folded intermediate under native conditions. In addition, we observed that amide protons located in the hinge between the two globular domains share similar 螖G_gu^app and m values and should unfold simultaneously. These observations provide new points of view for improving our understanding of the thermodynamic stability and the mechanisms that drive folding鈥搖nfolding dynamics of proteins.