J. Bukowska and P. Piotrowski
42
periments indicated that for the short thiol linkers the electron-transfer kinetics is
determined by protein dynamics rather than by electron tunneling probabilities and
that the reorientation of protein is controlled by the interfacial electric field. thus,
they show that SERRS can provide a very deep insight into dynamics of interfacial
processes of proteins on the molecular level. SERRS experiments deliver much
information about the heme part of the protein. A unique possibility of observing
vibrational spectrum of both the heme and amino acid residues is however offered
by tERS spectroscopy as demonstrated in 2008 [118].
Surface enhanced Raman scattering spectroscopy is now widely reported for
enzymes. In this case, combination of SERS and resonance Raman spectroscopy is
also usually applied, because it offers considerably higher sensitivity as compared
to SERS. Excellent review on the development of SERS for the detection of enzyme
transformations was published by Larmour et al. in 2010 [119]. Initially, SERRS
was used for direct detection of enzymes. First results on SERRS of glucose oxidase
has been reported at the very beginning of the SERS studies [120]. the first studies, in which biological activity of enzyme immobilized on Ag nanoparticles (Ag
colloid) was monitored by SERRS spectroscopy, were published in 1993 [121]. In
this report, 60–85 % retention of enzymatic activity of chlorocatechol dioxygenase
in the reaction of catechol substrate with oxygen was demonstrated. the subsequent
experiments for enzyme-substrate, enzyme-inhibitor, and enzyme-product complexes indicated that in general, SERRS allows probing the enzymatic processes
in situ, during catalytic turnover. however, it has to be stressed that in all SERRS
studies reported, some loss of the enzyme activity was observed upon contact with
the metal support responsible for signal enhancement. therefore, preparation of
biocompatible substrates seems to be a key problem and important challenge in
these experiments.
An alternative approach to study enzyme bioactivity with SERRS is recording the spectrum of enzymatically produced dyes, which are easily detected with
SERRS. For example, azo dyes may be used in such experiments because they exhibit strong and very characteristic SERRS spectrum. SERRS signal of azoaniline
adsorbed on Au nanoparticles has been applied for determination of glucose concentration [122], or adsorbed on Ag colloids for detection of antigen (mouse Igg)
[123]. In this case azo dye was produced in peroxidase catalyzed oxidation reaction
of o-phenylenediamine by hydrogen peroxide (Fig. 3.7).
the idea of utilizing SERRS spectrum of enzymatically produced dyes to detect
enzymes was further developed by Stevenson et al. [124]. In a very interesting
experiment, the SERRS spectrum of oxidized form of 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABtS), used as a substrate in an enzyme-linked immunosorbent assay (ELISA), was applied for the indirect detection of human prostate
specific antigen (PSA) [124]. ABtS yields a green product upon enzymatic reaction with horseradish peroxidase and is commonly used as a colorimetric indicator
of peroxidase activity. Experiments by Stevenson et al. [124] proved that SERRS
provides a lower limit of detection and a wider range of linearity of the Raman
signal vs. concentration as compared to the standard colorimetric ELISA approach.
42
periments indicated that for the short thiol linkers the electron-transfer kinetics is
determined by protein dynamics rather than by electron tunneling probabilities and
that the reorientation of protein is controlled by the interfacial electric field. thus,
they show that SERRS can provide a very deep insight into dynamics of interfacial
processes of proteins on the molecular level. SERRS experiments deliver much
information about the heme part of the protein. A unique possibility of observing
vibrational spectrum of both the heme and amino acid residues is however offered
by tERS spectroscopy as demonstrated in 2008 [118].
Surface enhanced Raman scattering spectroscopy is now widely reported for
enzymes. In this case, combination of SERS and resonance Raman spectroscopy is
also usually applied, because it offers considerably higher sensitivity as compared
to SERS. Excellent review on the development of SERS for the detection of enzyme
transformations was published by Larmour et al. in 2010 [119]. Initially, SERRS
was used for direct detection of enzymes. First results on SERRS of glucose oxidase
has been reported at the very beginning of the SERS studies [120]. the first studies, in which biological activity of enzyme immobilized on Ag nanoparticles (Ag
colloid) was monitored by SERRS spectroscopy, were published in 1993 [121]. In
this report, 60–85 % retention of enzymatic activity of chlorocatechol dioxygenase
in the reaction of catechol substrate with oxygen was demonstrated. the subsequent
experiments for enzyme-substrate, enzyme-inhibitor, and enzyme-product complexes indicated that in general, SERRS allows probing the enzymatic processes
in situ, during catalytic turnover. however, it has to be stressed that in all SERRS
studies reported, some loss of the enzyme activity was observed upon contact with
the metal support responsible for signal enhancement. therefore, preparation of
biocompatible substrates seems to be a key problem and important challenge in
these experiments.
An alternative approach to study enzyme bioactivity with SERRS is recording the spectrum of enzymatically produced dyes, which are easily detected with
SERRS. For example, azo dyes may be used in such experiments because they exhibit strong and very characteristic SERRS spectrum. SERRS signal of azoaniline
adsorbed on Au nanoparticles has been applied for determination of glucose concentration [122], or adsorbed on Ag colloids for detection of antigen (mouse Igg)
[123]. In this case azo dye was produced in peroxidase catalyzed oxidation reaction
of o-phenylenediamine by hydrogen peroxide (Fig. 3.7).
the idea of utilizing SERRS spectrum of enzymatically produced dyes to detect
enzymes was further developed by Stevenson et al. [124]. In a very interesting
experiment, the SERRS spectrum of oxidized form of 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABtS), used as a substrate in an enzyme-linked immunosorbent assay (ELISA), was applied for the indirect detection of human prostate
specific antigen (PSA) [124]. ABtS yields a green product upon enzymatic reaction with horseradish peroxidase and is commonly used as a colorimetric indicator
of peroxidase activity. Experiments by Stevenson et al. [124] proved that SERRS
provides a lower limit of detection and a wider range of linearity of the Raman
signal vs. concentration as compared to the standard colorimetric ELISA approach.