424
K. malek et al.
widely used in studies on protein secondary structure since both of these are sensitive to structural configuration. these vibrations assigned predominantly to the
C = o stretching and Nh bending modes, respectively, are observed in the 1670–
1620 and 1560–1530 cm
−1
regions. other bands characteristic for proteins only give
maxima in the high-wavenumber region above 3020 cm
−1
and they are attributed
to the Nh stretching vibrations (amide A and B). however, amide A and B are not
usually treated as spectral biomarkers. the amide I mode is most extensively used
to quantify α-helical, β-sheet, turn, and unordered structures of proteins. Usually,
IR spectra of cells and tissues exhibit amide bands at ca. 1665 and 1540 cm
−1
that
signify protein α-helical chains. The appearance of shoulders at ca. 1695–1670 and
1640–1620 cm
−1
indicates, in turn, changes in protein secondary conformation towards a β-sheet structure. For instance, such shifting was observed in IR spectra
of red blood cells from β-thalassemia patients [13] and during spontaneous differentiation of murine embryonic stem cells in their early development [14], while
β-amyloid peptides were detected in Alzheimer’s diseased tissue [15]. on the other
hand, the appearance of a band at 1624–1630 cm
−1
was also correlated with the
protein nitration process in the atherosclerotic lesion formation [16]. Shifting of the
amide I band should be carefully interpreted, because dispersion artefacts can appear in this spectral range. In such cases, the wavenumber value of this band can be
randomly shifted ca. 15 cm
−1
and its absorbance is often reduced [17].
A number of less pronounced bands from proteins appear in the region of 1460–
1240 cm
−1
, however, vibrations of other biomolecules can contribute to these bands
(see table 15.1 for details). In the region of the scissoring Ch 3 mode, a band at
1397 cm
−1
is assigned to proteins with some contribution from lipids, whereas a
band at 1400 cm
−1
is attributed to the symmetric stretching mode of the free carboxylate group of amino acids as well as fatty acids. In some cases, IR bands highlight
the presence of proteins containing tyr and Cys residues. the breathing vibration
of the tyrosine ring appears at ca. 1520 cm
−1
, while the stretching mode of the
cysteine thiol group is assigned to a weak and broad band with a maximum around
2550 cm
−1
. Both of these are rarely observed in raw IR spectra but are observed in
the second derivative spectra [13, 18]. In turn, the stretching C-oh mode of serine,
threonine, and tyrosine were found at 1171 cm
−1
in FtIR spectra of malignant oesophageal and neoplastic cervical tissues due to the absence of the glycogen band
normally observed at 1155 cm
−1
in normal tissue [19, 20].
FtIR spectra of some tissue sections exhibit markers characteristic of connective
tissue. this tissue mainly consists of proteins (e.g. collagen) with a small component of nucleic acids. A typical feature for the connective tissue is a triad of absorbances in the region of the amide III vibrations, i.e. 1206, 1238, and 1280 cm
−1
originating mainly from collagen. It is easily recognized by the positions of the
peaks and the relative intensities, which are accompanied by a broad amide II band
at 1550 cm
−1
[21]. In turn, cells in malignant tissue often exhibit an increase in the
intensity of the 970–965 cm
−1
band, which is generally correlated to an increase in
the relative content of nucleic acids. however, phosphorylation of newly formed
proteins, for example, in the process of carcinogenesis [19] can also contribute to
this band. A similar band was also observed when applying synchrotron IR mea-
K. malek et al.
widely used in studies on protein secondary structure since both of these are sensitive to structural configuration. these vibrations assigned predominantly to the
C = o stretching and Nh bending modes, respectively, are observed in the 1670–
1620 and 1560–1530 cm
−1
regions. other bands characteristic for proteins only give
maxima in the high-wavenumber region above 3020 cm
−1
and they are attributed
to the Nh stretching vibrations (amide A and B). however, amide A and B are not
usually treated as spectral biomarkers. the amide I mode is most extensively used
to quantify α-helical, β-sheet, turn, and unordered structures of proteins. Usually,
IR spectra of cells and tissues exhibit amide bands at ca. 1665 and 1540 cm
−1
that
signify protein α-helical chains. The appearance of shoulders at ca. 1695–1670 and
1640–1620 cm
−1
indicates, in turn, changes in protein secondary conformation towards a β-sheet structure. For instance, such shifting was observed in IR spectra
of red blood cells from β-thalassemia patients [13] and during spontaneous differentiation of murine embryonic stem cells in their early development [14], while
β-amyloid peptides were detected in Alzheimer’s diseased tissue [15]. on the other
hand, the appearance of a band at 1624–1630 cm
−1
was also correlated with the
protein nitration process in the atherosclerotic lesion formation [16]. Shifting of the
amide I band should be carefully interpreted, because dispersion artefacts can appear in this spectral range. In such cases, the wavenumber value of this band can be
randomly shifted ca. 15 cm
−1
and its absorbance is often reduced [17].
A number of less pronounced bands from proteins appear in the region of 1460–
1240 cm
−1
, however, vibrations of other biomolecules can contribute to these bands
(see table 15.1 for details). In the region of the scissoring Ch 3 mode, a band at
1397 cm
−1
is assigned to proteins with some contribution from lipids, whereas a
band at 1400 cm
−1
is attributed to the symmetric stretching mode of the free carboxylate group of amino acids as well as fatty acids. In some cases, IR bands highlight
the presence of proteins containing tyr and Cys residues. the breathing vibration
of the tyrosine ring appears at ca. 1520 cm
−1
, while the stretching mode of the
cysteine thiol group is assigned to a weak and broad band with a maximum around
2550 cm
−1
. Both of these are rarely observed in raw IR spectra but are observed in
the second derivative spectra [13, 18]. In turn, the stretching C-oh mode of serine,
threonine, and tyrosine were found at 1171 cm
−1
in FtIR spectra of malignant oesophageal and neoplastic cervical tissues due to the absence of the glycogen band
normally observed at 1155 cm
−1
in normal tissue [19, 20].
FtIR spectra of some tissue sections exhibit markers characteristic of connective
tissue. this tissue mainly consists of proteins (e.g. collagen) with a small component of nucleic acids. A typical feature for the connective tissue is a triad of absorbances in the region of the amide III vibrations, i.e. 1206, 1238, and 1280 cm
−1
originating mainly from collagen. It is easily recognized by the positions of the
peaks and the relative intensities, which are accompanied by a broad amide II band
at 1550 cm
−1
[21]. In turn, cells in malignant tissue often exhibit an increase in the
intensity of the 970–965 cm
−1
band, which is generally correlated to an increase in
the relative content of nucleic acids. however, phosphorylation of newly formed
proteins, for example, in the process of carcinogenesis [19] can also contribute to
this band. A similar band was also observed when applying synchrotron IR mea-