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B. Czarnik-Matusewicz and Y.M. Jung
first stage, i.e., within the initial 6 min, amide protons from the structures most
exposed to solvent, the outer surface of hAS, are exchanged for deuterons. this
process was identified from the changes at approximately 1,683 cm
−1
, which were
assigned to the β-turns. During the next stage, between 6 and 14 min, because of the
penetration of d 2 o deeper into the more compact interior of the hSA, protons from
the β-turns that were initially less solvent accessible are exchanged and give rise to
the peak at ~ 1,666 cm
−1
. In the last part of the h/d exchange, the peak observed at
1,653 cm
−1
shows marked proton replacement in the helical part that forms 67 % of
the total structure of HSA. It was estimated that approximately 25 % of the α-helix
structures in hSA are unaffected by the h/d exchange even 4 hours after the beginning of the experiment.
the time evolution of the spectral variations induced in the amide I and amide I’
range by h-d exchange at different ph values has been applied to examine the flexibility of immunoglobulin; this flexibility is essential for the function of Igg [76].
In the analysis, the heterocorrelation variant was also employed to confirm the correlations between two separate data sets measured at different ph values. the h/d
perturbation-based 2dCoS results have delivered evidence that Igg is composed
of multiple regions with differential flexibility and conformational heterogeneity;
these regions were differentially perturbed by solution conditions such as the ph.
the h/d-perturbed 2dCoS has allowed for an improved understanding of the relationships between Igg dynamics and stability.
the secondary and tertiary structures of the truncated protein covering the first
120 aa of the hepatitis C virus core protein (hCvc-120), which is assembled in
vitro into nucleocapsid-like particles, have also been studied through the dynamics
of h/d exchange [77]. the infrared spectra measured as a function of the h/d exchange time, over a period of two hours, were examined by both principal component analysis and two-dimensional correlation spectroscopy. the PCA results were
a good basis for the selection of two time domains for further two-dimensional correlation analysis. For PC1, the spectra collected over the initial 10 min were clearly
distinct from those measured between 13 and 120 min after the initiation of the h/d
isotopic exchange. the temporal characteristics of the h/d exchange obtained from
the 2d analysis performed separately for the two sets of data has shown that in most
arginine residues, protons from side-chain fragment were exchanged within the
first 10 min. this finding has allowed the postulate that the positively charged Arg
residues, which are easily accessible to aqueous environment, were prone to accept
negatively charged phosphate groups from the viral RNA during the assembly of the
hCv; this process should have led to an increase in the thermodynamic stability of
the nucleocapsid. the fast h/d exchange process for the arginine residues were followed by changes in the β-turns and unordered structures and were finally followed
by rearrangements in the β-sheets, which might have linked the protein monomers.
h/d exchange supported by simulation of this process has been a common procedure used by Pastrana-Rios’s group. For example, this approach has allowed the
resolution of the dynamics of the molecular events that occur upon h/d exchange
in two of eight different epidermal growth factor receptor (EGFR) ligands (TGF-α
and EgF) [71]. the studies were motivated by the unfortunate large overlap be-
B. Czarnik-Matusewicz and Y.M. Jung
first stage, i.e., within the initial 6 min, amide protons from the structures most
exposed to solvent, the outer surface of hAS, are exchanged for deuterons. this
process was identified from the changes at approximately 1,683 cm
−1
, which were
assigned to the β-turns. During the next stage, between 6 and 14 min, because of the
penetration of d 2 o deeper into the more compact interior of the hSA, protons from
the β-turns that were initially less solvent accessible are exchanged and give rise to
the peak at ~ 1,666 cm
−1
. In the last part of the h/d exchange, the peak observed at
1,653 cm
−1
shows marked proton replacement in the helical part that forms 67 % of
the total structure of HSA. It was estimated that approximately 25 % of the α-helix
structures in hSA are unaffected by the h/d exchange even 4 hours after the beginning of the experiment.
the time evolution of the spectral variations induced in the amide I and amide I’
range by h-d exchange at different ph values has been applied to examine the flexibility of immunoglobulin; this flexibility is essential for the function of Igg [76].
In the analysis, the heterocorrelation variant was also employed to confirm the correlations between two separate data sets measured at different ph values. the h/d
perturbation-based 2dCoS results have delivered evidence that Igg is composed
of multiple regions with differential flexibility and conformational heterogeneity;
these regions were differentially perturbed by solution conditions such as the ph.
the h/d-perturbed 2dCoS has allowed for an improved understanding of the relationships between Igg dynamics and stability.
the secondary and tertiary structures of the truncated protein covering the first
120 aa of the hepatitis C virus core protein (hCvc-120), which is assembled in
vitro into nucleocapsid-like particles, have also been studied through the dynamics
of h/d exchange [77]. the infrared spectra measured as a function of the h/d exchange time, over a period of two hours, were examined by both principal component analysis and two-dimensional correlation spectroscopy. the PCA results were
a good basis for the selection of two time domains for further two-dimensional correlation analysis. For PC1, the spectra collected over the initial 10 min were clearly
distinct from those measured between 13 and 120 min after the initiation of the h/d
isotopic exchange. the temporal characteristics of the h/d exchange obtained from
the 2d analysis performed separately for the two sets of data has shown that in most
arginine residues, protons from side-chain fragment were exchanged within the
first 10 min. this finding has allowed the postulate that the positively charged Arg
residues, which are easily accessible to aqueous environment, were prone to accept
negatively charged phosphate groups from the viral RNA during the assembly of the
hCv; this process should have led to an increase in the thermodynamic stability of
the nucleocapsid. the fast h/d exchange process for the arginine residues were followed by changes in the β-turns and unordered structures and were finally followed
by rearrangements in the β-sheets, which might have linked the protein monomers.
h/d exchange supported by simulation of this process has been a common procedure used by Pastrana-Rios’s group. For example, this approach has allowed the
resolution of the dynamics of the molecular events that occur upon h/d exchange
in two of eight different epidermal growth factor receptor (EGFR) ligands (TGF-α
and EgF) [71]. the studies were motivated by the unfortunate large overlap be-