Analyzing the Interaction of ADF/Cofilin with Actin through Molecular Dynamics Simulations

Actin is a cytoskeletal protein found in all eukaryotic cells. It is involved in cellular motility and cytokinesis and is essential for processes such as defense against microorganisms and for cancer cells to spread as metastases. Actin binds ATP in physiological conditions and hydrolyzes it to ADP after polymerizing into microfilaments. ADF/cofilin is one of the key actin-binding proteins and is required for the rapid actin turnover necessary in cell motility. It functions by taking advantage of the ATP/ADP gradient in actin filaments by associating with the older parts of actin filaments to depolymerize filaments by binding with 100 fold higher affinity to ADP-actin than to ATP-actin. ADF/cofilin also binds monomeric ADP-actin more strongly than ATP-actin and inhibits nucleotide exchange. However, the structural mechanisms for these processes are a mystery as a crystal structure is not available for the actin-cofilin complex. Yet, ADF/cofilin binds actin through an ADF homology (ADF-H) domain, which is also found in other actin-binding proteins, such as twinfilin, and a structure of the complex of ATP-actin with the C-terminal ADF-H domain of twinfilin was recently determined. We are using the crystal structure of the actin-Twf-C complex to simulate the interaction of ADF/cofilin with ADP-actin and ATP-actin through molecular dynamics simulations and to examine conformational changes that may explain the mechanisms for ADF/cofilin inhibition of nucleotide exchange as well as the stronger affinity for ADP-actin. Thus far we have run a 2 ns simulation of the Twf-C-actin complex with ATP. Surprisingly, in this simulation the adenine ring of ATP underwent a large ring flip even though in simulations of monomeric ATP-actin the ATP is extremely stable. This result was unexpected since a similar ring flip was previously seen in ADP-Arp3 (actin-related protein 3), but in this case the nucleotide was being released. Experimentally, however, ADF/cofilin prevents nucleotide release. It may be possible that the ATP is actually going into a more stable conformation. We are working on running the simulation for a longer time as well as replacing ADP into the complex. Analyzing ADF/cofilin’s contributions to cell motility through computational biology may help us understand its roles in apoptosis and several diseases, such as Alzheimer’s and cancer.