Crowders-Induced Compaction of Multi-kbp Long DNA Molecules Followed by TPM

Abstract

DNA organization in bacteria contributes to their environmental adaptation and survival. This includes the DNA compaction induced by a wide variety of nucleoproteins binding to DNA, especially the nucleoid associated proteins, as well as the presence of high concentrations of nonbinding crowders. Weak multivalent interactions and depletion effects, which are central to this process of DNA compaction, can be affected by the experimental constraints. Here, we combine experiments and polymer physics theory to quantitatively determine to what extent the Tethered Particle Motion can faithfully monitor the conformational changes of DNA molecules up to 15 kbp in length, resulting from their immersion in a medium crowded with synthetic and biological molecules, namely, PEG and BSA. By developing a theoretical model that includes monomer–monomer interactions modified by the depletion interaction due to the presence of crowders as well as the ones due to the substrate and the TPM particle, we show that the reversible coil–globule transition induced by crowders can be safely observed by TPM with only a slight effect of the presence of the substrate and the particle. The experimental results are consistent with previous bulk experiments and well-fitted by our detailed theoretical model, which also shows that the transition crowder concentration decreases with the length of the DNA molecule and proportionally to the size of the crowders. DNA compaction can therefore be studied by TPM in the near future with nucleoid associated proteins.