444 — Electrochemical analysis of polynucleotides

Abstract

Natural and synthetic polynucleotides were studied by means of differential and normal pulse polarography (d.p.p., n.p.p.) and by cyclic voltammetry (c.v.) in connection with a d.m.e. or h.m.d.e.oaIt was shown that the d.p.p. behaviour of synthetic double-stranded polydeoxyribonucleotides is markedly influenced by the nucleotide sequence; homopolymer pairs (poly(dA) · poly(dT) and poly(dG) · poly(dC)) yield almost no signal under the given conditions, whereas polynucleotides with the alternating nucleotide sequence (poly(dA-dT) · poly(dA-dT) and poly(dG-dC) · poly(dG-dC)) produce well-developed peaks. This difference may be connected with differences in the secondary structures of the two types of polynucleotides.bIt was demonstrated that both synthetic and natural polynucleotides containing guanine produce a characteristic anodic c.v. peak at about −0.2 V (vs s.c.e.). It is necessary to polarize the h.m.d.e. to potentials of about −1.8 V in order to obtain this peak. Polynucleotides which do not contain guanine (poly(A) · poly(C) and poly(U)) do not yield this anodic peak. Similar to cathodic responses, the anodic peak is substantially higher in denatured (single-stranded) DNA than in its native double-helical form.cVarious polynucleotides were modified with osmium tetroxide in an attempt to introduce a new electroactive centre into the polynucleotide chain. It is known that OsO4 reacts with pyrimidine bases in single-stranded polynucleotides. It does not react, however, with bases incorporated in an intact Watson-Crick DNA double helix; this agent can thus act as a chemical probe of DNA structure. It was found that the introduction of osmium into singlestranded poly(C), poly(U), poly(dT) and denatured DNA resulted in the appearance of several reduction signals. It was shown that the d.p.p. behaviour of synthetic double-stranded polydeoxyribonucleotides is markedly influenced by the nucleotide sequence; homopolymer pairs (poly(dA) · poly(dT) and poly(dG) · poly(dC)) yield almost no signal under the given conditions, whereas polynucleotides with the alternating nucleotide sequence (poly(dA-dT) · poly(dA-dT) and poly(dG-dC) · poly(dG-dC)) produce well-developed peaks. This difference may be connected with differences in the secondary structures of the two types of polynucleotides. It was demonstrated that both synthetic and natural polynucleotides containing guanine produce a characteristic anodic c.v. peak at about −0.2 V (vs s.c.e.). It is necessary to polarize the h.m.d.e. to potentials of about −1.8 V in order to obtain this peak. Polynucleotides which do not contain guanine (poly(A) · poly(C) and poly(U)) do not yield this anodic peak. Similar to cathodic responses, the anodic peak is substantially higher in denatured (single-stranded) DNA than in its native double-helical form. Various polynucleotides were modified with osmium tetroxide in an attempt to introduce a new electroactive centre into the polynucleotide chain. It is known that OsO4 reacts with pyrimidine bases in single-stranded polynucleotides. It does not react, however, with bases incorporated in an intact Watson-Crick DNA double helix; this agent can thus act as a chemical probe of DNA structure. It was found that the introduction of osmium into singlestranded poly(C), poly(U), poly(dT) and denatured DNA resulted in the appearance of several reduction signals. After the treatment of native (double-helical) DNA with OsO4 this DNA yielded d.p.p. peaks which are substantially smaller than peaks of single-stranded polynucleotides treated in the same manner. The peaks of the modified native DNA increased if DNA was damaged by various agents such as ionizing and U.V. radiations, which introduce minor local changes into the double helix of DNA.