Direct evidence for the presence of left-handed conformation in a supramolecular assembly of polynucleotides.
Hexammine cobalt(III) chloride (Co(NH3)6(3+) provokes a B-DNA----Z-DNA----psi-DNA conformational transition in poly(dG-dC).poly(dG-dC) and poly(dG-m5dC).poly(dG-m5dC). The circular dichroism spectrum of psi-DNA is characterized by a manyfold increase of positive ellipticity in the range of 300-225 nm and the complete absence of a negative peak. In order to ascertain the helical handedness of psi-DNA, we used a recently developed enzyme immunoassay technique. This method consisted of treating the polynucleotides with Co(NH3)6(3+) to convert them to the Z- or psi-DNA forms and immobilizing these conformations on a microtiter plate. The plates were subsequently treated with a monoclonal anti-Z-DNA antibody Z22, alkaline phosphatase conjugated, affinity purified immunoglobulins, and the phosphatase substrate. The enzyme-substrate reaction was monitored by reading the absorbance at 405 nm with a microplate autoreader. The monoclonal anti-Z-DNA antibody had no reactivity to the B-DNA form, but bound strongly to both the Z- and psi-DNA forms, showing that Co(NH3)6(3+)-induced psi-DNA form of the polynucleotides exists in the left-handed Z-DNA conformation.
Thomas
T J
TJ
Department of Medicine, UMDNJ--Robert Wood Johnson Medical School, New Brunswick 08903.
Thomas
T
T
D000906:Antibodies; D000911:Antibodies, Monoclonal; D002942:Circular Dichroism; D003035:Cobalt; D004247:DNA; D007124:Immunoenzyme Techniques; D009690:Nucleic Acid Conformation; D011089:Polydeoxyribonucleotides
D000906:Antibodies; D000911:Antibodies, Monoclonal; D011089:Polydeoxyribonucleotides; C030862:cobalt ammonium complex; D003035:Cobalt; C029988:poly(dG-m(5)dC).poly(dG-m(5)dC); C033927:poly(dA-dC).poly(dG-dT); D004247:DNA