SYNTHESIS OF A DNA KNOT CONTAINING BOTH POSITIVE AND NEGATIVE NODES

A figure-fight (4(1)) knot, containing two positive and two negative nodes, has been synthesized by ligating a single-stranded DNA molecule into a closed cycle. The molecule consists of two single-turn double-helical domains connected by four dT14 or dT15 linkers. One of the domains contains the sequence (dCpdGp)6, Which is capable of forming left-handed Z-DNA. Ligation yields a trefoil (3(1)) knot or a 4(1) knot, depending upon the conditions in solution. A 4(1) knot results if Co(NH3)63+ is present in the solution to promote the formation of Z-DNA; a 3(1) knot is the product in the absence of Co(NH3)63+. Both domains of the 3(1) knot are susceptible to restriction endonuclease digestion, but only the domain of the 4(1) knot expected to be right-handed can be digested by a restriction endonuclease. The 4(1) knot has both a higher gel-electrophoretic mobility at low polyacrylamide concentrations and a higher friction constant than the 3(1) knot, when analyzed by Ferguson plots under denaturing conditions.