analysis of in-vivo lacr-mediated gene repression based on the mechanics of dna looping分析已lacr-mediated基因镇压基于dna循环的机制.pdfVIP

Analysis of In-Vivo LacR-Mediated Gene Repression Based on the Mechanics of DNA Looping 1¤ 2 1 2,3 Yongli Zhang *, Abbye E. McEwen , Donald M. Crothers , Stephen D. Levene * 1 Departments of Chemistry and Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut, United States of America, 2 Institute of Biomedical Sciences and Technology, University of Texas at Dallas, Richardson, Texas, United States of America, 3 Department of Molecular and Cell Biology, University of Texas at Dallas, Richardson, Texas, United States of America Interactions of E. coli lac repressor (LacR) with a pair of operator sites on the same DNA molecule can lead to the formation of looped nucleoprotein complexes both in vitro and in vivo. As a major paradigm for loop-mediated gene regulation, parameters such as operator affinity and spacing, repressor concentration, and DNA bending induced by specific or non-specific DNA- binding proteins (e.g., HU), have been examined extensively. However, a complete and rigorous model that integrates all of these aspects in a systematic and quantitative treatment of experimental data has not been available. Applying our recent statistical-mechanical theory for DNA looping, we calculated repression as a function of operator spacing (58–156 bp) from first principles and obtained excellent agreement with independent sets of in-vivo data. The results suggest that a linear extended, as opposed to a closed v-shaped, LacR conformation is the dominant form of the tetramer in vivo. Moreover, loop- mediated repression in wild-type E. coli strains is facilitated by decreased DNA rigidity and high levels of flexibility in the LacR tetramer. In contrast, repression data for strains lacking HU gave a near-normal value of the DNA persistence length. These finding