A temperature-regulated iso-hyperbaric spectrophotometer: construction and performance characteristics.
The position of an equilibrium is a function of pressure and temperature. Generally, only temperature effects are discussed for biochemical systems; however, pressure perturbations can offer interesting insights into the role of hydration in a reaction. We describe the construction and characteristics of an instrument that allows computer control of the temperature and pressure throughout the range -40 to +140 degrees C and 0.1 to 250 MPa, as well as automatic data acquisition from either a spectrophotometer or a spectrofluorimeter. To test the performance of the system the effect of pressure on the thermal denaturation of poly[d(A-T)] and poly(dA).poly(dT) has been investigated. In 20 mM NaCl, 20 mM Tris-HCl, pH 8.9, the delta V degrees for poly[d(A-T)] equals +0.30 +/- 0.09 cm3/mol (base pairs), and for poly(dA).poly(dT) delta V degrees equals +3.05 +/- 0.15 cm3/mol (base pairs). Although we focus on the use of this instrument to measure thermal denaturation curves, it can be employed to study the equilibrium of any reaction that can be followed optically in the ultraviolet or visible.
Wu
J Q
JQ
Department of Medicinal Chemistry, University of Illinois, Chicago 60612.
Macgregor
R B
RB
D004247:DNA / Q000737:chemistry; D004867:Equipment Design; D006874:Hydrostatic Pressure; D008838:Microcomputers; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D012984:Software; D013053:Spectrophotometry / Q000295:instrumentation*; D013696:Temperature; D013816:Thermodynamics
D004247:DNA