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1.
2.
Mikhail M. Vorobev 《Journal of Molecular Catalysis .B, Enzymatic》2009,58(1-4):146-152
Proteolysis of casein substrates includes demasking stage, the transition of masked bonds to the demasked stage, where peptide bonds become accessible to the enzyme attack. Therefore, proteolysis was regarded as a two-stage process with consequent demasking and hydrolysis stages. When demasking process is kinetically significant, the peptide bonds are hydrolysed with some lag. It was shown both by theoretical simulations and experimentally that the increase of amino nitrogen can be a non-monotonous function of the hydrolysis degree or proteolysis time. The non-monotonously dependence was found for chymotryptic proteolysis of β-casein, while for α-casein the monotonous dependence was obtained. This was treated as an indication of the prevalence of the hydrophobically induced masking effect for β-casein. For the proteolysis of β-casein by wild-type and engineered trypsins, the kinetic analysis allowed us to conclude that demasking stage was initiated by the splitting of the main peptide chain, which compact conformation was initially stabilized by the interaction of hydrophobic regions of peptide chain. 相似文献
3.
Ram Prasad Bora Mehmet Ozbil Rajeev Prabhakar 《Journal of biological inorganic chemistry》2010,15(4):485-495
In this B3LYP study, the catalytic mechanisms for the hydrolysis of the three different peptide bonds (Lys28-Gly29, Phe19-Phe20,
and His14-Gln15) of Alzheimer amyloid beta (Aβ) peptide by insulin-degrading enzyme (IDE) have been elucidated. For all these
peptides, the nature of the substrate was found to influence the structure of the active enzyme–substrate complex. The catalytic
mechanism is proposed to proceed through the following three steps: (1) activation of the metal-bound water molecule, (2)
formation of the gem-diol intermediate, and (3) cleavage of the peptide bond. With the computed barrier of 14.3, 18.8, and
22.3 kcal/mol for the Lys28-Gly29, Phe19-Phe20, and His14-Gln15 substrates, respectively, the process of water activation
was found to be the rate-determining step for all three substrates. The computed energetics show that IDE is the most efficient
in hydrolyzing the Lys28-Gly29 (basic polar–neutral nonpolar) peptide bond followed by the Phe19-Phe20 (neutral nonpolar–neutral
nonpolar) and His14-Gln15 (basic polar–neutral polar) bonds of the Aβ substrate. 相似文献
4.
In recent years, many advances have been made in the understanding of functional and structural characteristics of protein
evolution from denaturant-based studies that subject the protein to a change in the microenvironment. This paper reports the
chemical denaturation of purified goat muscle cystatin (GMC) a thiol-proteinase inhibitor, using urea and guanidine hydrochloride
(GdnHCl). The subtle conformational changes of GMC were monitored by intrinsic fluorescence, extrinsic fluorescence, and CD
spectroscopic techniques. Further, the activity of GMC as a function of increasing concentration of denaturants was also studied.
It was found that increasing the concentration of GdnHCl significantly enhances the inactivation and unfolding of the inhibitor
(GMC). In urea-induced denaturation, the intrinsic and extrinsic fluorescence intensity reveals significant structural changes
in the inhibitor. Further, it was found that at low concentrations of urea, up to 0.5–1.0 M, there was quenching of fluorescence intensity compared with the native form and a red shift of 5 nm was observed up to 5–8 M. The results presented in this paper suggest that GdnHCl-induced denaturation of GMC follows a simple two-state rule in which
native → denatured state transition occurs in a single step. However denaturation with urea proceeds through an intermediate
or non-native state. 相似文献
5.
Gurusamy Balakrishnan Mohammed Ibrahim Piotr J. Mak Jessica Hata James R. Kincaid Thomas G. Spiro 《Journal of biological inorganic chemistry》2009,14(5):741-750
Time-resolved resonance Raman (RR) spectra are reported for hemoglobin (Hb) tetramers, in which the α and β chains are selectively
substituted with mesoheme. The Soret absorption band shift in mesoheme relative to protoheme permits chain-selective recording
of heme RR spectra. The evolution of these spectra following HbCO photolysis shows that the geminate recombination rates and
the yields are the same for the two chains, consistent with recent results on 15N-heme isotopomer hybrids. The spectra also reveal systematic shifts in the deoxyheme ν
4 and ν
Fe–His RR bands, which are anticorrelated. These shifts are resolved for the successive intermediates in the protein structure,
which have previously been determined from time-resolved UV RR spectra. Both chains show Fe–His bond compression in the immediate
photoproduct, which relaxes during the formation of the first intermediate, Rdeoxy (0.07 μs), in which the proximal F-helix is proposed to move away from the heme. Subsequently, the Fe–His bond weakens, more
so for the α chains than for the β chains. The weakening is gradual for the β chains, but is abrupt for the α chains, coinciding
with completion of the R–T quaternary transition, at 20 μs. Since the transition from fast- to slow-rebinding Hb also occurs
at 20 μs, the drop in the α chain ν
Fe–His supports the localization of ligation restraint to tension in the Fe–His bond, at least in the α chains. The mechanism is
more complex in the β chains. 相似文献
6.
Ajikumar P. K. Devaky K. S. 《International journal of peptide research and therapeutics》2000,7(4):207-215
Summary Studies leading to optimization of butanediol dimethacrylate-crosslinked polystyrene supports (BDDMA-PS) for solid phase peptide
synthesis are delineated. BDDMA-PS copolymers with different crosslink densities were prepared and functionalised with chloromethyl
groups. The reactivity of the Lys(2-Cl−Z)−OH residue bound to these polymers through a benzyl ester linkage was investigated
by following the kinetics of acylation by the HOBt active ester of Boc-Alanine. From the results it was observed that the
rate of peptide bond formation was maximum for a 2% BDDMA crosslinked resin. This resin was compared with a 2% DVB-crosslinked
polystyrene resin (DVB-PS). Synthesis of an extremely insoluble, hydrophobic, antiparallel β-sheeted difficult sequence peptide
LMVGGVVIA (β 34–42), C-terminal fragment of β-amyloid protein, β (1–42), was carried out on both 2% DVB-PS and 2% BDDMA-crosslinked
polystyrene supports. The synthesis of the peptide was carried out using Boc amino acid strategy. Greater extent of swelling
of the resino peptide, increased coupling efficiency during the assembly of amino acids and relatively high purity of synthesised
peptide were observed in the case of 2% BDDMA-PS polymer. 相似文献
7.
Emilia Bramanti Francesco Lenci Antonella Sgarbossa 《European biophysics journal : EBJ》2010,39(11):1493-1501
We have determined the secondary structure of 1–40 β-amyloid peptides by Fourier-transform infrared spectroscopy (FTIR) and
characterized the peptide photophysical properties before and after self-assembly by using intrinsic tyrosine steady-state
and time-resolved fluorescence. All measurements were performed in the presence and absence of hypericin (Hyp), an exogenous
natural polycyclic pigment that has been shown to inhibit fibril formation and has also been used as a fluorescent probe.
We monitored the time course of the aggregation process measuring 405 nm light diffusion at 90° and used thioflavin T to reveal
the presence of fibrils. FTIR quantitative analysis evidenced a prevalent random conformation at t = 0 with and without Hyp. Fibrils showed a predominant parallel β-sheet structure and a small percentage of α-helix. The
results of fluorescence measurements showed that Hyp does significantly interact with peptides in β-sheet conformation. In
conclusion, hypericin does hinder the formation of fibrils, but the percentages of parallel β-sheets were not significantly
different from those found in samples not treated with Hyp. 相似文献
8.
Accumulating evidence suggests that Aβ1–42–membrane interactions may play an important role in the pathogenesis of Alzheimer’s disease. However, the mechanism of this
structural transition remains unknown. In this work, we have shown that submicellar concentrations of sodium dodecyl sulfate
(SDS) can provide a minimal platform for Aβ1–42 self-assembly. To further investigate the relation between Aβ1–42 structure and function, we analyzed peptide conformation and aggregation at various SDS concentrations using circular dichroism
(CD), Fourier transform infrared spectroscopy, and gel electrophoresis. These aggregates, as observed via atomic force microscopy,
appeared as globular particles in submicellar SDS with diameters of 35–60 nm. Upon sonication, these particles increased in
disc diameter to 100 nm. Pyrene I
3/I
1 ratios and 1-anilinonaphthalene-8-sulfonic acid binding studies indicated that the peptide interior is more hydrophobic than
the SDS micelle interior. We have also used Forster resonance energy transfer between N-terminal labeled pyrene and tyrosine
(10) of Aβ1–42 in various SDS concentrations for conformational analysis. The results demonstrate that SDS at submicellar concentrations
accelerates the formation of spherical aggregates, which act as niduses to form large spherical aggregates upon sonication.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
9.
Ken’ichiroh Nakamura Hiroaki Mori Toru Kawakami Hironobu Hojo Yoshiaki Nakahara Saburo Aimoto 《International journal of peptide research and therapeutics》2007,13(1-2):191-202
The 4,5-dimethoxy-2-mercaptobenzyl (Dmmb) group attached to a main chain amide in a peptide is easily transformed into an
S-peptide via an intramolecular N–S acyl shift reaction under acidic conditions, and the S-peptide produces a peptide thioester through an intermolecular thiol–thioester exchange reaction. In order to develop a method
for efficiently preparing peptide thioesters based on the N–S acyl shift reaction, the factors involved in this process were analyzed in detail. The general features of the transformation
at the Dmmb group attached amide bond in a trifluoroacetic acid (TFA) solution and the generation of a peptide thioester were
examined by 13C-NMR spectral measurements, reversed-phase (RP) HPLC analyses, mass measurements, and amino acid analyses. The methoxy group
of the Dmmb group was not essential for the N–S acyl shift reaction, but played a role in stabilizing the thioester form. The addition of water to the TFA solution accelerated
the N–S acyl shift reaction mediated by the Dmmb group and also suppressed the acid-catalyzed cleavage of the Dmmb group. A peptide
thioester was produced from the S-peptide via an intermolecular thiol–thioester exchange reaction with minimal epimerization of the amino acid residue that constituted
the thioester bond. Undesirable side reactions, such as the hydrolysis of the thioester bond and an S–N acyl shift reaction occurred during the synthetic process, which is a subject of further investigation. 相似文献
10.
Gehman JD O'Brien CC Shabanpoor F Wade JD Separovic F 《European biophysics journal : EBJ》2008,37(3):333-344
Aβ(1–42) peptide, found as aggregated species in Alzheimer’s disease brain, is linked to the onset of dementia. We detail
results of 31P and 2H solid-state NMR studies of model membranes with Aβ peptides and the effect of metal ions (Cu2+ and Zn2+), which are found concentrated in amyloid plaques. The effects on the lipid bilayer and the peptide structure are different
for membrane incorporated or associated peptides. Copper ions alone destabilise the lipid bilayer and induce formation of
smaller vesicles, but not when Aβ(1–42) is associated with the bilayer membrane. Aβ(25–35), a fragment from the C-terminal
end of Aβ(1–42), which lacks the metal coordinating sites found in the full length peptide, is neurotoxic to cortical cortex
cell cultures. Addition of metal ions has little effect on membrane bilayers with Aβ(25–35) peptides. 31P magic angle spinning NMR data show that Aβ(1–42) and Aβ(1–42)-Cu2+ complexes interact at the surface of anionic phospholipid membranes. Incorporated peptides, however, appear to disrupt the
membrane more severely than associated peptides. Solid-state 13C NMR was used to compare structural changes of Aβ(1–42) to those of Aβ(25–35) in model membrane systems of anionic phospholipids
and cholesterol. The Aβ peptides appeared to have an increase in β-strand structure at the C-terminus when added to phospholipid
liposomes. The inclusion of Cu2+ also influenced the observed chemical shift of residues from the C-terminal half, providing structural clues for the lipid-associated
Aβ/metal complex. The results point to the complex pathway(s) for toxicity of the full-length peptide.
Australian Society for Biophysics Special Issue: Metals and Membranes in Neuroscience. 相似文献
11.
Polyakova O Dear D Stern I Martin S Hirst E Bawumia S Nash A Dodson G Bronstein I Bayley PM 《European biophysics journal : EBJ》2009,38(2):209-218
Formation of PrP aggregates is considered to be a characteristic event in the pathogenesis of TSE diseases, accompanied by
brain inflammation and neurodegeneration. Factors identified as contributing to aggregate formation are of interest as potential
therapeutic targets. We report that in vitro proteolysis of ovine PrP94–233 (at neutral pH and in the absence of denaturants) by the protease cathepsin S, a cellular enzyme that also shows enhanced
expression in pathogenic conditions, occurs selectively in the region 135–156. This results in an unusually efficient, concentration-dependent
conformational conversion of a large subfragment of PrP94–233 into a soluble β-structured oligomeric intermediate species, that readily forms a thioflavin-T-positive aggregate. N-terminal
sequencing of the proteolysis fragments shows the aggregating species have marked sequence similarities to truncated PrP variants
known to confer unusually severe pathogenicity when transgenically expressed in PrPo/o mice. Circular dichroism analysis shows that PrP fragments 138–233, 144–233 and 156–233 are significantly less stable than
PrP94–233. This implies an important structural contribution of the β1 sequence within the globular domain of PrP. We propose that
the removal or detachment of the β1 sequence enhances β-oligomer formation from the globular domain, leading to aggregation.
The cellular implications are that specific proteases may have an important role in the generation of membrane-bound, potentially
toxic, β-oligomeric PrP species in pre-amyloid states of prion diseases. Such species may induce cell death by lysis, and
also contribute to the transport of PrP to neuronal targets with subsequent amplification of pathogenic effects.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
12.
Chartchalerm Isarankura-Na-Ayudhya Tanawut Tantimongcolwat Hans-Joachim Galla Virapong Prachayasittikul 《Biological trace element research》2010,134(3):352-363
In the present study, spectroscopic determinations of copper ions using chimeric metal-binding green fluorescent protein (His6GFP)
as an active indicator have been explored. Supplementation of copper ions to the GFP solution led to a remarkable decrease
of fluorescent intensity corresponding to metal concentrations. For circumstances, rapid declining of fluorescence up to 60%
was detected in the presence of 500 μM copper. This is in contrast to those observed in the case of zinc and calcium ions,
in which approximately 10–20% of fluorescence was affected. Recovery of its original fluorescence up to 80% was mediated by
the addition of ethylenediamine tetraacetic acid. More importantly, in the presence of metal ions, the emission wavelength
maximum remains unchanged while reduction of the optical density of the absorption spectrum has been observed. This indicates
that the chromophore’s ground state was possibly affected by the static quenching process. Results from circular dichroism
measurements revealed that the overall patterns of circular dichroism spectra after exposure to copper ions were not significantly
different from that of the control, where the majority of sharp positive band around 195–196 nm in combination with a broad
negative deflection around 215–216 nm was obtained. Taken together, it can be presumed that copper ions exerted their static
quenching on the fluorescence rather than structural or conformational alteration. However, notification has to be made that
some peptide rearrangements may also occur in the presence of metal ions. Further studies were conducted to investigate the
feasibility of using the His6GFP as a sensing unit for copper ions. The His6GFP was encapsulated in Sol-gel and immobilized
onto the optical fiber connected with a fluorescence detecting device. The Sol-gel was doped into the metal solution where
the quenching of fluorescence could be monitored in real time. The sensing unit provided a high sensitivity of detection in
the range of 0.5 μM to 50 mM with high selectivity for copper ions. All these findings open up a high potential to apply the
fluorescent protein-based bioanalytical tool for copper determination in the future. 相似文献
13.
Protein composition and native state of chlorophylls were analyzed in two wheat (Triticum durum L.) genotypes with different tolerance to drought, Barakatli-95 (drought-tolerant) and Garagylchyg-2 (drought-sensitive),
during water deficit. It is shown that the plants subjected to water deficit appear to have a slight increase in α-and β-subunits
of CF1ATP-synthase complex (57.5 and 55 kD, respectively) in Barakatli-95 and their lower content in Garagylchyg-2. Steady-state
levels of the core antenna of PS II (CP47 and CP43) and light-harvesting Chl a/b-apoproteins (LHC) II in the 29.5–24 kD region remained more or less unchanged in both wheat genotypes. The synthesis of 36
kD protein and content of low-molecular-weight polypeptides (21.5, 16.5, and 14 kD) were noticeably increased in the tolerant
genotype Barakatli-95. Drought caused significant changes in the carotenoid region of the spectrum (400–500 nm) in drought-sensitive
genotype Garagylchyg-2 (especially in the content of pigments of the violaxanthin cycle). A shift of the main band from 740–742
to 738 nm is observed in the fluorescence spectra (77 K) of chloroplasts from both genotypes under water deficiency, and there
is a stimulation of the ratio of fluorescence band intensity F687/F740.
Published in Russian in Biokhimiya, 2006, Vol. 71, No. 2, pp. 223–228. 相似文献
14.
Subfragments of amyloid-beta (Aβ) appear to protect neurons from Alzheimer’s disease (AD). The permeability of the blood–brain
barrier (BBB) has limited in vivo research. The aim of this study is to explore permeation of the BBB by chitosan nanoparticles loaded with Aβ and to evaluate
immunogenicity of these particles. Chitosan microspheres were prepared by mechanical stirring emulsification methods combined
with chemical crosslinking. Morphological characteristics of the nanoparticles were examined using high-resolution transmission
electron microscopy. The peptide association efficiency was determined by high-performance liquid chromatography. Fluorescently
labeled chitosan nanoparticle-intramembranous fragments of Aβ (NP-IF-A) were administered systemically to mice in order to
evaluate brain translocation by fluorescence microscopy. The immunogenicity of the nano-vaccine was determined by enzyme-linked
immunosorbent assay (ELISA). All nanoparticles analyzed were well-separated, roughly spherical structures with uniform particle
size distribution in the range of 15.23 ± 10.97 nm. The peptide association efficiency was 78.4%. The brain uptake efficiency
of nano-antigen was 80.6%; uptake efficiency of antigen alone was only 20.6%. ELISA showed that the nano-vaccine had favorable
immunogenicity. A chitosan nano-carrier for Aβ allowed permeation of the BBB and was non-immunogenic. These findings indicate
that this novel targeted nano-vaccine delivery system can be used as a carrier for Aβ. This system will further research of
peptide vaccines for AD. 相似文献
15.
Yuan C Yi L Yang Z Deng Q Huang Y Li H Gao Z 《Journal of biological inorganic chemistry》2012,17(2):197-207
Amyloid beta (Aβ) peptide accumulation has been demonstrated to play a central role in Alzheimer’s disease (AD). Substantial
evidence indicates that protein nitrotyrosination contributes to Aβ-dependent neurotoxicity; however, the molecular mechanism
is unknown. Recent research has shown that Aβ complexes with heme to form Aβ–heme, and increases the pseudo-peroxidase activity
of heme. We found that Aβ–heme uses H2O2 and NO2
− to cause nitration of enolase and synaptic proteins more effectively than heme. Thus, the increased peroxidase activity of
Aβ–heme may be the molecular link between excess Aβ and the widespread protein nitration in AD. Interestingly, the site of
enolase nitration that was catalyzed by Aβ–heme is different from that induced by heme. Moreover, the secondary structural
perturbations of Aβ–heme-treated and heme-treated enolase are also different. These observations suggest that Aβ–heme targets
specific amino acid sequences in enolase. Furthermore, our data show that Aβ–heme peroxidase activity is independent of the
aggregation state of Aβ, suggesting an important role of soluble Aβ in addition to Aβ aggregates and oligomers in AD pathogenesis. 相似文献
16.
Lakshman PL Tachibana S Toyama H Taira T Suganuma T Suntornsuk W Yasuda M 《Journal of industrial microbiology & biotechnology》2011,38(9):1485-1492
An acid proteinase from Monascus purpureus No. 3403, MpuAP, was previously purified and some characterized in our laboratory (Agric Biol Chem 48:1637–1639, 1984). However, further information about this enzyme is lacking. In this study, we investigated MpuAP’s comprehensive substrate
specificity, storage stability, and prospects for reducing antigenicity of whey proteins for application in the food industry.
MpuAP hydrolyzed primarily five peptide bonds, Gln4–His5, His10–Leu11, Ala14–Leu15, Gly23–Phe24 and Phe24–Phe25 in the oxidized insulin B-chain. The lyophilized form of the enzyme was well preserved at 30–40°C for 7 days without stabilizers.
To investigate the possibility of reducing the antigenicity of the milk whey protein, enzymatic hydrolysates of the whey protein
were evaluated by inhibition ELISA. Out of the three main components of whey protein, casein and α-lactalbumin were efficiently
degraded by MpuAP. The sequential reaction of MpuAP and trypsin against the whey protein successfully degraded casein, α-lactalbumin
and β-lactoglobulin with the highest degree of hydrolysis. As a result, the hydrolysates obtained by using the MpuAP–trypsin
combination showed the lowest antigenicity compared with the single application of pepsin, trypsin or pepsin–trypsin combination.
Therefore, the overall result suggested that the storage-stable MpuAP and trypsin combination will be a productive approach
for making hypoallergic bovine milk whey protein hydrolysates. 相似文献
17.
Fanyan Meng Chuanlai Shen Fengqin Miao Yong He Yuqing Shen Wei Xie Jianqiong Zhang 《Biotechnology letters》2010,32(12):1803-1808
The heavy chain protein of HLA-peptide complexes (HLA/HBc18–27 and HLA/CEA694–702) immobilized onto an ion exchange chromatography column and then the dilution-refolded HBc18–27-fused or CEA694–702-fused β2m protein was able to pass through the column. Using this method, HLA/peptide complexes were prepared within 30 h
with a refolding yield of at least 20% (w/w) and purity of over 80% (w/w). This strategy refolds, concentrates, and purifies
HLA/peptide complexes in a single integrated step and offers a potential tool to refold multiple-subunit proteins other than
the major histocompatibility complex (MHC)/peptide complexes. 相似文献
18.
David J. Wilton Ryo Kitahara Kazuyuki Akasaka Mike P. Williamson 《Journal of biomolecular NMR》2009,44(1):25-33
Pressure-dependent 13C chemical shifts have been measured for aliphatic carbons in barnase and Protein G. Up to 200 MPa (2 kbar), most shift changes
are linear, demonstrating pressure-independent compressibilities. CH3, CH2 and CH carbon shifts change on average by +0.23, −0.09 and −0.18 ppm, respectively, due to a combination of bond shortening
and changes in bond angles, the latter matching one explanation for the γ-gauche effect. In addition, there is a residue-specific component, arising from both local compression and conformational
change. To assess the relative magnitudes of these effects, residue-specific shift changes for protein G were converted into
structural restraints and used to calculate the change in structure with pressure, using a genetic algorithm to convert shift
changes into dihedral angle restraints. The results demonstrate that residual 13Cα shifts are dominated by dihedral angle changes and can be used to calculate structural change, whereas 13Cβ shifts retain significant dependence on local compression, making them less useful as structural restraints. 相似文献
19.
Steer David L. Lew Rebecca A. Perlmutter Patrick Smith A. Ian Aguilar Marie-Isabel 《International journal of peptide research and therapeutics》2001,8(3-5):241-246
Summary The use of β-amino acids as peptidomimetics has emerged in recent years with significant potential in a number of applications.
The incorporation of β-amino acids has been successful in creating peptidomimetics that not only have potent biological activity,
but are also resistant to proteolysis. This article reviews the recent applications of β-amino acids in the design of protease
and peptidase inhibitors. Given their structural diversity, together with the ease of synthesis and incorporation into peptide
sequences using standard solid-phase peptide synthesis techniques, β-amino acids have the potential to form a new platform
technology for peptidomimetic design and synthesis. 相似文献
20.
Hugo M. Botelho Sónia S. Leal Andreas Veith Vesna Prosinecki Christian Bauer Renate Fröhlich Arnulf Kletzin Cláudio M. Gomes 《Journal of biological inorganic chemistry》2010,15(2):271-281
Rieske proteins and Rieske ferredoxins are present in the three domains of life and are involved in a variety of cellular
processes. Despite their functional diversity, these small Fe–S proteins contain a highly conserved all-β fold, which harbors
a [2Fe–2S] Rieske center. We have identified a novel subtype of Rieske ferredoxins present in hyperthermophilic archaea, in
which a two-cysteine conserved SKTPCX(2–3)C motif is found at the C-terminus. We establish that in the Acidianus ambivalens representative, Rieske ferredoxin 2 (RFd2), these cysteines form a novel disulfide bond within the Rieske fold, which can
be selectively broken under mild reducing conditions insufficient to reduce the [2Fe–2S] cluster or affect the secondary structure
of the protein, as shown by visible circular dichroism, absorption, and attenuated total reflection Fourier transform IR spectroscopies.
RFd2 presents all the EPR, visible absorption, and visible circular dichroism spectroscopic features of the [2Fe–2S] Rieske
center. The cluster has a redox potential of +48 mV (25 °C and pH 7) and a pK
a of 10.1 ± 0.2. These shift to +77 mV and 8.9 ± 0.3, respectively, upon reduction of the disulfide. RFd2 has a melting temperature
near the boiling point of water (T
m = 99 °C, pH 7.0), but it becomes destabilized upon disulfide reduction (ΔT
m = −9 °C, ΔC
m = −0.7 M guanidinium hydrochloride). This example illustrates how the incorporation of an additional structural element such
as a disulfide bond in a highly conserved fold such as that of the Rieske domain may fine-tune the protein for a particular
function or for increased stability. 相似文献