Cystine, an essential determinant of protein tertiary structure, is also a target for electrochemical manipulation: Cyclotides (original) (raw)
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Peptides represent a promising class of biorecognition elements that can be coupled to electrochemical transducers. The benefits lie mainly in their stability and selectivity toward a target analyte. Furthermore, they can be synthesized rather easily and modified with specific functional groups, thus making them suitable for the development of novel architectures for biosensing platforms, as well as alternative labelling tools. Peptides have also been proposed as antibiofouling agents. Indeed, biofouling caused by the accumulation of biomolecules on electrode surfaces is one of the major issues and challenges to be addressed in the practical application of electrochemical biosensors. In this review, we summarise trends from the last three years in the design and development of electrochemical biosensors using synthetic peptides. The different roles of peptides in the design of electrochemical biosensors are described. The main procedures of selection and synthesis are discussed. Sel...
A Sensitive Electrochemical Protein Quantification Method
Electroanalysis, 2000
This article presents a sensitive electrochemical method for the quanti®cation of proteins. The assay used the biuret reaction, which is based on the complexation of proteins with copper ions to form a copper-protein complex. In the present approach, once copper ions were complexed with the protein, they were released by an acidic treatment and simultaneously separated by ultracentrifugation from the protein sample. This allowed differential pulse anodic stripping voltammetry (DPASV) of Cu 2 on carbon rotogravure printed electrodes (CARPEs). A nM detection limit for bovine serum albumin (BSA) was found with this method. The kinetics of this assay were tested and it resulted that the total assay can be carried out in 27 min, including incubation, washing steps and detection.
Recent developments in capillary and microchip electroseparations of peptides (2011-2013)
ELECTROPHORESIS, 2013
Recent developments in capillary and microchip electroseparations of peptides (2011-2013) * The review presents a comprehensive survey of recent developments and applications of capillary and microchip electroseparation methods (zone electrophoresis, ITP, IEF, affinity electrophoresis, EKC, and electrochromatography) for analysis, isolation, purification, and physicochemical and biochemical characterization of peptides. Advances in the investigation of electromigration properties of peptides, in the methodology of their analysis, including sample preseparation, preconcentration and derivatization, adsorption suppression and EOF control, as well as in detection of peptides, are presented. New developments in particular CE and CEC modes are reported and several types of their applications to peptide analysis are described: conventional qualitative and quantitative analysis, determination in complex (bio)matrices, monitoring of chemical and enzymatical reactions and physical changes, amino acid, sequence and chiral analysis, and peptide mapping of proteins. Some micropreparative peptide separations are shown and capabilities of CE and CEC techniques to provide relevant physicochemical characteristics of peptides are demonstrated.
Electrochemical detection and characterization of proteins
Biosensors and …, 2006
On-line detection of serum proteins is of clinical relevance, in detecting leaks and biofouling in hemofiltration equipment, biofilm growth on prosthetic devices, or hemolysis within a prosthetic or therapeutic device. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were employed to detect and analyze micromolar concentrations of four globular proteins of clinical importance. CV testing showed that identification and quantification of each of these proteins was possible through analysis of current changes at specific potentials. Preliminary CV studies into the contamination of Bovine Serum Albumin with a microgram amount of one of the other three proteins illustrated that direct detection of the contaminant protein was possible. The analysis of the EIS data demonstrated that with increase in relative concentration of proteins, the amount of electroactive proteins adsorption at the interface increases, leading to increase in surface charge density and capacitance, especially for lower molecular weight proteins. The impedance data was used to determine the values of Gibbs adsorption energy, adsorption coefficients for the four proteins, and develop an equivalent circuit model for the protein-containing solutions.
Journal of Electroanalytical Chemistry, 2016
A novel bio-electrochemical immunosensor for sensitive and selective detection of anti-peptide antibodies of HPV was fabricated by immobilization of a peptide (SPINNTKPHEAR) (1) linked to a 6-aminohexanoic (Ahx) residue and ferrocene (Fc) on a gold electrode surface, used as sensing interface. The Fc-Ahx-peptide (Fc-Ahx-SPINNTKPHEAR) (1AFc) was obtained by solid phase peptide synthesis by using the Fmoc/tBu strategy. Peptides were purified by preparative HPLC, characterized by MS, MALDI-TOF, and Circular Dichroism and their purity was evaluated by using analytical HPLC. The electrochemical behavior of the modified electrode was examined by cyclic voltammetry in phosphate buffer solution (PBS) in order to evaluate the redox behavior of the ferrocene moiety. The influence of anti-peptide antibodies on the voltammetric response of the modified electrode was investigated by comparing results obtained with pre-immune (control) and post immune serum samples at similar dilution factor. Changes in such behavior upon addition of the serum samples to PBS suggested that the fabricated bioelectrochemical sensor was able to recognize the interaction between anti-peptide antibodies and the immobilized peptide (1AFc) with high selectivity and sensitivity. Such influence increased as the dilution decreased, but the effect was less pronounced at relatively high concentrated solutions owing to the effect of the sample matrix. Notwithstanding, the proposed biosensor can clearly detect the target anti-peptide antibodies at a significant large concentration range.
Journal of Chromatography B, 2003
Boron-doped diamond thin film (BDD) electrodes have been used to study the oxidation reactions and to detect leucine-enkephalinamide (LEA) and its metabolites, tyrosine (T), tyrosyl-alanine (TA), tyrosyl-alanine-glycine (TAG) and leucine-enkephalin (LE) using cyclic voltammetry (CV), flow-injection analysis (FIA), and gradient liquid chromatography (LC) with amperometric detection. At diamond electrodes, well-defined and highly reproducible cyclic voltammograms were obtained with signal-to-background (S /B) ratios 5-10 times higher than those observed for glassy carbon (GC) electrodes. The analytical peaks of LC for LEA and its metabolites were well resolved. No deactivation of BDD electrodes was found after several experiments with standard as well as plasma samples, indicating high stability of the electrode. Calibration curves were linear over a wide range from 0.06 to 30 mM with regression coefficients of 0.999 for all compounds. The limits of detection obtained based on a signal-to-noise ratio of 3:1 were 3, 2.2, 2.7, 20 and 11 nM for T, TA, TAG, LE and LEA, respectively. These values were at least one order lower than those obtained at GC electrodes, which has given limits of detection of 22.88, 20.64, 89.57, 116.04 and 75.67 for T, TA, TAG, LE and LEA, respectively. Application of this method to real samples was demonstrated and validated using rabbit serum samples. This work shows the promising use of conducting diamond as an amperometric detector in gradient LC, especially for the analysis of enkephalinamide and its metabolites.
A glassy carbon electrode,covered with a Nafion film, on which nano-structured bimetallic particles of Ag-Hg were deposited; from now on "the AgHgNpNf/GC electrode", was used for the indirect determination of cysteine, glutathione and metallothioneins by measuring the catalytic hydrogen evolution signal according to the Brdička reaction using cobalt as a catalyst. Upon successful formation and deposition of the Ag-Hg bimetallic nanoparticles cyclic voltammetry (CV), scanning electron microscopy (SEM) and atomic force microscopy (AFM) were used to characterize the electrode for that specific task. The results showed the electrode to be mechanically resistant even under sonication, stable in its performance, porous, sensitive in its response and of low cost to be built. Differential pulse voltammetry signals for cysteine and glutathione obtained using the AgHgNpNf/GC electrode were qualitatively similar to those reported using a hanging mercury drop electrode, (HMDE), dropping mercury electrode (DME) and static mercury drop electrode (SMDE). Detection limits based on the variability of a blank solution, (3 s criterion) calculated for those tests were 0.463 g L −1 and 0.064 g L −1 , for cysteine and glutathione, respectively. Based on the Brdička procedure and considering the particular characteristics of the new electrode, some analytical tests were carried out for the indirect determination of the metallothioneins content of samples of blood serum of rats exposed to lead by measuring the evolution of the catalytic hydrogen signal. An average accuracy value around 102%, n = 6, and a precision of 0.87% ± 0.09 were obtained for these tests.
Microchemical Journal, 2000
Ž. 3q Ru bpy-based chemiluminescence detection of underivatized amino acids was coupled to capillary electrophore-3 sis and evaluated as a separation and detection methodology for the compositional analysis of peptides and proteins. Two tripeptides were studied: glycine᎐phenylalanine᎐alanine, and valine᎐proline᎐leucine, and the method was demonstrated to be quantitative and reproducible at the 10-g and 5-g peptide levels, with a limit of detection Ž. Ž. Ž. 3q LOD of 2.5 pmol for gly᎐phe᎐ala and 80 fmol for val᎐pro᎐leu SrNs 2. Ru bpy-based chemiluminescence 3 detection coupled to CE offers a sensitive and time-efficient method for the determination of the amino acid composition of peptides or proteins without derivatization. The detection limits for most amino acids were demonstrated to be at, or below the level achievable using ninhydrin or phenyl isothiocyanate derivatization. Amino acids were identified by their migration time through the capillary and their relative luminescence. The relative Ž. 3q luminescent response obtained upon reaction of Ru bpy with amino acids was dependent upon the amino acid 3 R-group at the ␣-carbon, and the relative response may be useful in broadly classifying a number of amino acids.
Electroanalysis, 1998
ABSTRACT Constant current derivative chronopotentiometric stripping analysis (CPSA) was used to study bioactive peptides [Lys8]-vasopressin and angiotensin II at carbon paste electrode (CPE) and hanging mercury drop electrode (HMDE). Both peptides contain a single tyrosine residue which is oxidized at CPE close to +0.6 V (against Ag/AgCl/3 M KCl electrode) producing peak Y; this peak was by about 15 mV more positive in vasopressin than in angiotensin II. At HMDE vasopressin yielded peak S due to the reduction of the disulfidic bond in the peptide. No peak S was observed with angiotensin II as it does not contain any cystine/cysteine residues. Vasopressin produced different catalytic hydrogen reduction signals in media with cobalt ion and in media not containing any metal ion. The signal produced in the latter media (peak H), which appeared close to −1.7 V, was well separated from the background. No catalytic hydrogen reduction signal was obtained with angiotensin II. The above mentioned CPSA signals can be applied for the determination of the respective peptide at nanomolar concentrations. The least sensitive peak S appears which requires at least 100 nM vasopressin concentration at moderate accumulation time. By two orders of magnitude, higher sensitivities can be obtained with peaks Y and H. In measuring these peaks CPSA shows significant advantages over the voltammetric stripping techniques.