Influence of DNA acidification on DNA premelting and template properties

Area of Science:

• Molecular Biology
• Biophysics

Background:

• DNA conformation can be altered by pH changes.
• Such alterations may affect DNA-protein interactions and biological functions.

Purpose of the Study:

• To investigate the effects of acidification on T7 DNA conformation.
• To determine how these conformational changes impact RNA polymerase binding and transcription.

Main Methods:

• UV difference spectroscopy at sub-melting temperatures.
• Analysis of poly(purine)-poly(pyrimidine) polymers.
• Assessing Escherichia coli RNA polymerase binding and transcription of acidified T7 DNA.

Main Results:

• Acidification induced partly irreversible conformational changes in T7 DNA.
• Poly(purine)-poly(pyrimidine) polymers showed altered premelting behavior after acidification.
• Acidified and reneutralized T7 DNA exhibited inhibited RNA synthesis and chain initiation.
• A decrease in the number and stability of RNA-polymerase-DNA complexes was observed.

Conclusions:

• Acidification leads to subtle, irreversible DNA conformational changes.
• These changes negatively affect RNA polymerase binding and transcription initiation.
• The stability and number of RNA-polymerase-DNA complexes are reduced.