hexamminecobalt(iii)-induced condensation of calf thymus DNA: circular dichroism and hydration measurements
B.I. Kankiar, V. Buckin and V. A. Bloomfield
Physicochemical and Engineering Aspects, 183-185, pp. 517-526 (2001)
The interaction of hexamminecobalt(lII), Co(NH3)6 3+, with 160 and 3000—8000 bp length calf thymus DNA has been investigated by circular dichroism, acoustic and densimetric techniques. The acoustic titration curves of 160 bp DNA revealed three stages of interaction: (i) Co(NH3)6 3+ binding up to the molar ratio [Co(NH3)6 3+]/[P] = 0.25, prior to DNA condensation; (ii) a condensation process between [Co(NH3)6 3+]/[P] = 0.25 and 0.30; and (iii) precipitation after [Co(NH3)6 3+]/[P] = 0.3. In the case of 3000— 8000 bp DNA only two processes were observed: (i) binding up to [Co(NH33)6 3+]/[P] = 0.3; and (ii) precipitation after this point. In agreement with earlier observations, long DNA aggregates without changes in its B-form circular dichroism spectrum, while short DNA demonstrates a positive B→φtransition after [Co(NH3)6 3+]/[P] = 0.25. From ultrasonic and densimetric measurements the effect of Co(NH3)6 3+ binding volume and compressibility have been obtained. The binding of Co(NH3)6 3+ to both short and long DNA is characterized by similar changes in volume and compressibility calculated per mole Co(NH3)6 3+: ΔV = 9 cm3 mol-1 and ΔK= 33 x 10-4 cm3 mol-1 bar-1. The positive sign of the parameters indicates dehydration, ie water release from Co(NH3)6 3+ and the atomic groups of DNA. This extent of water displacement would be consistent with the formation of two direct, hydrogen bonded contacts between the cation and the phosphates of DNA. DNA in viruses and cells exists in a highly condensed, tightly packed state. For instance, the concentration of DNA in the head of bacteriphages is ~800 mg/ml and in metaphase chromosomes ~170 mg/ml (1). In contrast, DNA concentration in spectroscopic studies is usually ~10-2 mg/ml. Knowledge of the condensation process is essential for understanding the molecular mechanisms of biological processes such as DNA transcription and replication. Studies on DNA condensation have received additional impetus in recent years from an interest in gene therapy, which is based on delivery of foreign DNA molecules into cells (2 and references therein).