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Correlation spectroscopy and molecular dynamics simulations to study the structural features of proteins.

Varriale A, Marabotti A, Mei G, Staiano M, D'Auria S - PLoS ONE (2013)

Bottom Line: Our results showed that keeping temperature constant, the protein diffusion coefficient decreased from 84±4 µm(2)/s to 44±3 µm(2)/s when pH was changed from 7.0 to 4.0.An even more marked decrease of the MalE2 diffusion coefficient (31±3 µm(2)/s) was registered when pH was raised from 7.0 to 10.0.The obtained fluorescence correlation data, corroborated by circular dichroism, fluorescence emission and light-scattering experiments, are discussed together with the MD simulations results.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Molecular Sensing, IBP-CNR, Naples, Italy. a.varriale@ibp.cnr.it

ABSTRACT
In this work, we used a combination of fluorescence correlation spectroscopy (FCS) and molecular dynamics (MD) simulation methodologies to acquire structural information on pH-induced unfolding of the maltotriose-binding protein from Thermus thermophilus (MalE2). FCS has emerged as a powerful technique for characterizing the dynamics of molecules and it is, in fact, used to study molecular diffusion on timescale of microsecond and longer. Our results showed that keeping temperature constant, the protein diffusion coefficient decreased from 84±4 µm(2)/s to 44±3 µm(2)/s when pH was changed from 7.0 to 4.0. An even more marked decrease of the MalE2 diffusion coefficient (31±3 µm(2)/s) was registered when pH was raised from 7.0 to 10.0. According to the size of MalE2 (a monomeric protein with a molecular weight of 43 kDa) as well as of its globular native shape, the values of 44 µm(2)/s and 31 µm(2)/s could be ascribed to deformations of the protein structure, which enhances its propensity to form aggregates at extreme pH values. The obtained fluorescence correlation data, corroborated by circular dichroism, fluorescence emission and light-scattering experiments, are discussed together with the MD simulations results.

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Dependence of diffusion coefficient (D) as a function of pH.The error bars of the measurements were obtained by using three different measurements. The histogram shows the aggregation percentage (axis on the right) as a function of pH values. The sphere and the two ellipsoids represent (in scale) the possible shapes of MalE2 (at pH 7.0, pH 4.0 and pH 10.0) compatible with the corresponding diffusion coefficients measured by FCS (see Table 1 and Materials and Methods section).
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pone-0064840-g002: Dependence of diffusion coefficient (D) as a function of pH.The error bars of the measurements were obtained by using three different measurements. The histogram shows the aggregation percentage (axis on the right) as a function of pH values. The sphere and the two ellipsoids represent (in scale) the possible shapes of MalE2 (at pH 7.0, pH 4.0 and pH 10.0) compatible with the corresponding diffusion coefficients measured by FCS (see Table 1 and Materials and Methods section).

Mentions: Autocorrelation spectra and residual distribution (inset) of MalE2 at pH 7.0 (panel A), pH 4.0 (panel B) and pH 10.0 (panel C). The best fitting of the data was obtained by the Equation 1.


Correlation spectroscopy and molecular dynamics simulations to study the structural features of proteins.

Varriale A, Marabotti A, Mei G, Staiano M, D'Auria S - PLoS ONE (2013)

Dependence of diffusion coefficient (D) as a function of pH.The error bars of the measurements were obtained by using three different measurements. The histogram shows the aggregation percentage (axis on the right) as a function of pH values. The sphere and the two ellipsoids represent (in scale) the possible shapes of MalE2 (at pH 7.0, pH 4.0 and pH 10.0) compatible with the corresponding diffusion coefficients measured by FCS (see Table 1 and Materials and Methods section).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3672191&req=5

pone-0064840-g002: Dependence of diffusion coefficient (D) as a function of pH.The error bars of the measurements were obtained by using three different measurements. The histogram shows the aggregation percentage (axis on the right) as a function of pH values. The sphere and the two ellipsoids represent (in scale) the possible shapes of MalE2 (at pH 7.0, pH 4.0 and pH 10.0) compatible with the corresponding diffusion coefficients measured by FCS (see Table 1 and Materials and Methods section).
Mentions: Autocorrelation spectra and residual distribution (inset) of MalE2 at pH 7.0 (panel A), pH 4.0 (panel B) and pH 10.0 (panel C). The best fitting of the data was obtained by the Equation 1.

Bottom Line: Our results showed that keeping temperature constant, the protein diffusion coefficient decreased from 84±4 µm(2)/s to 44±3 µm(2)/s when pH was changed from 7.0 to 4.0.An even more marked decrease of the MalE2 diffusion coefficient (31±3 µm(2)/s) was registered when pH was raised from 7.0 to 10.0.The obtained fluorescence correlation data, corroborated by circular dichroism, fluorescence emission and light-scattering experiments, are discussed together with the MD simulations results.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Molecular Sensing, IBP-CNR, Naples, Italy. a.varriale@ibp.cnr.it

ABSTRACT
In this work, we used a combination of fluorescence correlation spectroscopy (FCS) and molecular dynamics (MD) simulation methodologies to acquire structural information on pH-induced unfolding of the maltotriose-binding protein from Thermus thermophilus (MalE2). FCS has emerged as a powerful technique for characterizing the dynamics of molecules and it is, in fact, used to study molecular diffusion on timescale of microsecond and longer. Our results showed that keeping temperature constant, the protein diffusion coefficient decreased from 84±4 µm(2)/s to 44±3 µm(2)/s when pH was changed from 7.0 to 4.0. An even more marked decrease of the MalE2 diffusion coefficient (31±3 µm(2)/s) was registered when pH was raised from 7.0 to 10.0. According to the size of MalE2 (a monomeric protein with a molecular weight of 43 kDa) as well as of its globular native shape, the values of 44 µm(2)/s and 31 µm(2)/s could be ascribed to deformations of the protein structure, which enhances its propensity to form aggregates at extreme pH values. The obtained fluorescence correlation data, corroborated by circular dichroism, fluorescence emission and light-scattering experiments, are discussed together with the MD simulations results.

Show MeSH
Related in: MedlinePlus