Osmolyte mediation of T7 DNA polymerase and plasmid DNA stability.
The thermal stability of T7 DNA polymerase and pGEM4Z plasmid DNA in the presence and absence of the osmolyte N-methylglycine (sarcosine) was determined by means of UV spectroscopy. The decrease in melting temperature observed upon addition of sarcosine to solutions containing the plasmid DNA is linear with the concentration of sarcosine present. The enthalpy of the transition is also linear in its relationship to the melting temperature, and the entropy of the transition is linear in the natural log of the melting temperature. The slopes of both the entropic and enthalpic plots are equal. Destabilization of the plasmid DNA is observed to be entropically driven. The melting temperature of the T7 DNA polymerase complex is increased from 41 degrees C by addition of sarcosine to the solution. The relationship between the amount of sarcosine added and the melting temperature is linear, with a temperature of 61 degrees C observed for a 6 M solution. No clear trend of the effect of sarcosine on the enthalpy or entropy of the transitions could be observed.
Thakar
M
M
Department of Biochemistry, Ohio State University, Columbus 43210.
Bilenko
A
A
Becktel
W J
WJ
D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000378:metabolism*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D009995:Osmosis; D010957:Plasmids; D012521:Sarcosine / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet
D004247:DNA; C034970:bacteriophage T7 induced DNA polymerase; D004259:DNA-Directed DNA Polymerase; D012521:Sarcosine