Single molecule studies characterize the kinetic mechanism of tetrameric p53 binding to different native response elements

Abstract

The transcriptional activator p53 is a tumor suppressor protein that controls cellular pathways important for cell fate decisions, including cell cycle arrest, senescence, and apoptosis. It functions as a tetramer by binding to specific DNA sequences known as response elements (REs) to control transcription via interactions with co-regulatory complexes. Critical for understanding how p53 regulates gene expression is unraveling the fundamental mechanisms by which it binds to REs. Toward this goal we have used an in vitro single molecule fluorescence approach to quantify the dynamic binding of tetrameric p53 to different REs in real time under equilibrium conditions. For each RE tested we measured a single rate constant for association and two rate constants for dissociation, showing two kinetically distinguishable populations of tetrameric/RE complexes. Moreover, p53/DNA interactions were more dynamic as the RE sequence became closer to the consensus sequence. We propose a model in which p53 and its RE consensus sequence evolved to favor dynamic binding.