Molecular interaction mechanism of 14-3-3ε protein with phosphorylated Myeloid leukemia factor 1 revealed by an enhanced conformational sampling

An enhanced conformational sampling, a genetic-algorithm-based multidimensional virtual-system coupled molecular dynamics (GA-based mD-VcMD), was applied to a peptide, phosphorylated Myeloid leukemia factor 1 (pMLF1), binding to a receptor protein, 14-3-3e, in an explicit solvent. This method provides an equilibrated ensemble at room temperature, from which a density map of pMLF1 around 14-3-3e can be calculated. At a low-density level, pMLF1 distributed everywhere around 14-3-3e involving pMLF1 free from or slightly contacting to 14-3-3e. With increasing the density level, four clusters were identified on the 14-3-3e surface, and free-energy barriers existed among some clusters. At a further high-density level, a single cluster remained, which was characterized by the inter-molecular contacts commonly observed in the crystal structure, and the molecular orientation of pMLF1 was ordered as in the crystal structure. Increasing the density level, the cluster mentioned above was split into two high-density spots (semi-native-like and native-like spots) connected by a narrow corridor. When pMLF1 passing the corridor, salt-bridge relay (switching) regarding the phosphorylated residue of pMLF1 was occurred. The current study supports an amino acid mutagenesis experiment, which was conducted to measure the complex-structure stability. We also performed a simulation of non-phosphorylated MLF1, and showed that the complex structure was unstable, suggesting the important role of phosphorylation of MLF1 through activation of 14-3-3e in a variety of human diseases.