Metal binding induces conversion of B- to the hybrid B-Z-form in natural DNA.

Highly polymerized herring testis DNA of the random nucleotide sequence has been studied in solution by circular dichroism and ultra-violet absorption spectrometry under various experimental conditions. At low temperature upon addition of 0.05 M NaCl or 1.15 M MgSO4 the DNA formed a helix that belonged to the B-family. As the temperature was increased a transition from the pure B- to the hybrid B-Z-form occurred in the presence of 1.15 M MgSO4. This transition occurred over a large range of temperatures and corresponded to a non-cooperative conformational change. A similar DNA transition was induced with 0.098 mM Co(NH3)6Cl3. However, in the presence of 5.3 M NaCl the DNA conformation was not similar to that observed in 1.15 M MgSO4 or 0.098 mM Co(NH3)6Cl3 independently on the environmental temperature. In 5.3 M NaCl the DNA is thought to undergo a transition from one to another right-handed conformation that could be intermediate partially dehydrated conformer arising on the first step in the sequential transition to the dehydration of the polynucleotide. Our results show that a realistic model of native DNA, bearing Z-tracts embedded in B-helixes, can be obtained upon binding of alkaline earth or transition metals.

Action of hexaamminecobalt on the activity of Serratia marcescens nuclease.

Using CD spectroscopic and kinetic analysis, a refined mechanism of Co(NH3)6(3+) action on activity of Serratia marcescens nuclease was elucidated. The mechanism was identical with previously found mechanisms of Mg2+ and C7H5O2Hg+. Similarly to Mg2+ and C7H5O2Hg+, Co(NH3)6(3+) binding to the DNA substrate induced changes in the secondary structure which resulted in changes of the enzymatic activity of the S. marcescens nuclease. Upon binding of 0.03 Co(NH3)6(3+) per DNA phosphate, highly polymerized DNA displayed A-form characteristics. The DNA transition from B-form to A-form intermediate was followed by a decrease of the nuclease activity. The diminishing nuclease activity was consistent with diminishing values of Km and Kcat. Co(NH3)6(3+) binding to the highly polymerized DNA caused a 1.7-2.8-fold decrease in Km, and 13.3-19.9 decrease in Vmax compared with Mg-DNA complex. A vast excess of Co(NH3)6(3+) did not affect the activity of S. marcescens nuclease if the DNA in the assay mixture remained in its B-form conformation. Preincubation of S. marcescens nuclease with Co(NH3)6(3+) did not influence the tertiary structure of the enzyme.