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Albany 2001

category image Biomolecular
Stereodynamics
SUNY at Albany
June 19-23, 2001

Analysis of Peptides and Proteins With Electrodes

Electrochemical analysis with mercury and solid electrodes showed its usefulness in nucleic acid research (1-3). Recent progress in the development of electrochemical transducer-based devices for determining nucleotide sequences and measuring DNA damage is encouraging (4). The ability of peptides and proteins containing cysteine residues to produce catalytic electrochemical signals at mercury electrodes in cobalt containing solutions has been known for almost 70 years (5). The oxidation voltammetric signals obtained with carbon electrodes were little utilized in protein research because high concentrations of the biomacromolecules were necessary for their analysis (6). Recently we have shown (5, 7) that with the constant current chronopotentiometric stripping analysis (CPSA) remarkably low levels of peptides and proteins can be determined. CPSA of proteins is based on the oxidation of tyrosine and tryptophan residues in the protein molecule. Subnanogram amounts of bioactive peptides containing these amino acids can be analyzed. We have also shown that at mercury electrodes peptides and proteins produce "peak H" (5) at highly negative potentials (close to -1.7 V against SCE). Neither the presence of cysteine residues in the protein molecule nor cobalt ions in solution are necessary to obtain peak H. This peak is well developed and can be used for the determination of peptides and proteins at nanomolar concentrations (picogram amounts of peptides were sufficient for the analysis). Recently we applied the above electrochemical signals to study bioactive peptides and some proteins such as constructs of p53 protein, metallothioneins, histones, etc. We have found (8) that some osmium tetroxide complexes form adducts with proteins and peptides at neutral pH. These adducts produce several electrochemical signals. The most negative signal (at about -1.2 V) is due to catalytic hydrogen evolution and makes it possible to analyze osmium-modified peptides and proteins at low concentrations. Electrochemical techniques appear suitable not only for the determination of proteins but also for studies of nucleic acid-protein interactions and other biologically relevant purposes.

References and Footnotes
  1. Palecek, E. (1976) Prog .Nucleic Acid Res. Mol .Biol. 18, 151-213
  2. Palecek, E. (1983) in G. Milazzo (Ed.), Topics in Bioelectrochem. Bioenerg. Vol. 5, J. Wiley, Chichester, pp. 65-155
  3. Palecek, E. (1996) Electroanalysis 8, 7-14
  4. Palecek, E. and Fojta, M. (2001) Anal. Chem. 73, 75A-83A
  5. Tomschik, M., Havran, L., Fojta, M. and Palecek, E. (1998) Electroanalysis 10, 403-409
  6. Palecek, E., Jelen, F., Teijeiro, C., Fucik, V. and Jovin, T.M. (1993) Anal. Chim. Acta 273, 175-186
  7. Cai, X., Rivas, G., Farias, P. A. M., Shiraishi, H., Wang, J. and Palecek, E. (1996) Anal. Chim. Acta 332, 49-57
  8. Palecek, E., Billova, S., Kizek, R., unpublished

Emil Palecek, Sabina Billov?, Marie Br?zdov?, Ludek Havran, Rene Kizek and Mirek Tomschik

Institute of Biophysics, Academy of Sciences of the Czech Republic, Kr?lovopolsk? 135, 612 65 BRNO, Czech Republic
Phone: (4205) 74 62 41, 415 17 177 FAX:(4205) 41 21 12 93; email: palecek@ibp.cz