An innovative analytical/computational approach is presented to provide maximum allowed probabilities (MAPs) of conformations in protein domains not rigidly connected. The approach is applied to calmodulin and to its adduct with α-synuclein. Calmodulin is a protein'constituted by two rigid domains, each of them composed by two calcium-binding EF-hand motifs, which in solution are largely free to move with respect to one another. We used the N60D mutant of calmodulin, which had been engineered to selectively bind a paramagnetic lanthanide ion to only one of its four calcium binding sites, specifically in the second EF-hand motif of the N-terminal domain. In this way, pseudocontact shifts (pcs's) and self-orientation residual dipolar couplings (rdc's) measured on the C-terminal domain provide information on its relative mobility with respect to the domain hosting the paramagnetic center. Available NMR data for terbium(III) and thulium(III) calmodulin were supplemented with additional data for dysprosium(III), analogous data were generated for the α-synuclein adduct, and the conformations with the largest MAPs were obtained for both systems. The MAP analysis for calmodulin provides further information on the variety of conformations experienced by the system. Such variety is somewhat reduced in the calmodulin - α-synuclein adduct, which however still retains high flexibility. The flexibility of the calmodulin - α-synuclein adduct is an unexpected result of this research. © 2007 American Chemical Society.

Paramagnetism-based NMR restraints provide maximum allowed probabilities for the different conformations of partially independent protein domains / Bertini, Ivano; Gupta, Yogesh K.; Luchinat, Claudio; Parigi, Giacomo; Peana, Massimiliano; Sgheri, Luca; Yuan, Jing. - In: JOURNAL OF THE AMERICAN CHEMICAL SOCIETY. - ISSN 0002-7863. - 129:42(2007), pp. 12786-12794. [10.1021/ja0726613]

Paramagnetism-based NMR restraints provide maximum allowed probabilities for the different conformations of partially independent protein domains

PEANA, Massimiliano Francesco;
2007

Abstract

An innovative analytical/computational approach is presented to provide maximum allowed probabilities (MAPs) of conformations in protein domains not rigidly connected. The approach is applied to calmodulin and to its adduct with α-synuclein. Calmodulin is a protein'constituted by two rigid domains, each of them composed by two calcium-binding EF-hand motifs, which in solution are largely free to move with respect to one another. We used the N60D mutant of calmodulin, which had been engineered to selectively bind a paramagnetic lanthanide ion to only one of its four calcium binding sites, specifically in the second EF-hand motif of the N-terminal domain. In this way, pseudocontact shifts (pcs's) and self-orientation residual dipolar couplings (rdc's) measured on the C-terminal domain provide information on its relative mobility with respect to the domain hosting the paramagnetic center. Available NMR data for terbium(III) and thulium(III) calmodulin were supplemented with additional data for dysprosium(III), analogous data were generated for the α-synuclein adduct, and the conformations with the largest MAPs were obtained for both systems. The MAP analysis for calmodulin provides further information on the variety of conformations experienced by the system. Such variety is somewhat reduced in the calmodulin - α-synuclein adduct, which however still retains high flexibility. The flexibility of the calmodulin - α-synuclein adduct is an unexpected result of this research. © 2007 American Chemical Society.
Paramagnetism-based NMR restraints provide maximum allowed probabilities for the different conformations of partially independent protein domains / Bertini, Ivano; Gupta, Yogesh K.; Luchinat, Claudio; Parigi, Giacomo; Peana, Massimiliano; Sgheri, Luca; Yuan, Jing. - In: JOURNAL OF THE AMERICAN CHEMICAL SOCIETY. - ISSN 0002-7863. - 129:42(2007), pp. 12786-12794. [10.1021/ja0726613]
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11388/170644
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