Unraveling Complex Local Protein Environments with 4-Cyano-l-phenylalanine.

We present a multifaceted approach to effectively probe complex local protein environments utilizing the vibrational reporter unnatural amino acid (UAA) 4-cyano-l-phenylalanine (pCNPhe) in the model system superfolder green fluorescent protein (sfGFP). This approach combines temperature-dependent infrared (IR) spectroscopy, X-ray crystallography, and molecular dynamics (MD) simulations to provide a molecular interpretation of the local environment of the nitrile group in the protein. Specifically, a two-step enantioselective synthesis was developed that provided an 87% overall yield of pCNPhe in high purity without the need for chromatography.

Development and Validation of AMBER-FB15-Compatible Force Field Parameters for Phosphorylated Amino Acids.

Phosphorylation of select amino acid residues is one of the most common biological mechanisms for regulating protein structures and functions. While computational modeling can be used to explore the detailed structural changes associated with phosphorylation, most molecular mechanics force fields developed for the simulation of phosphoproteins have been noted to be inconsistent with experimental data. In this work, we parameterize force fields for the phosphorylated forms of the amino acids serine, threonine, and tyrosine using the ForceBalance software package with the goal of improving agreement with experiments for these residues.