The epidermal growth factor receptor (EGFR) is a member of the receptor tyrosine kinase family that plays a role in multiple cellular processes. Activation of EGFR requires binding of a ligand on the extracellular domain to promote conformational changes leading to dimerization and transphosphorylation of intracellular kinase domains. Seven ligands are known to bind EGFR with affinities ranging from sub-nanomolar to near micromolar dissociation constants. In the case of EGFR, distinct conformational states assumed upon binding a ligand is thought to be a determining factor in activation of a downstream signaling network. Previous biochemical studies suggest the existence of both low affinity and high affinity EGFR ligands. While these studies have identified functional effects of ligand binding, high-resolution structural data are lacking. To gain a better understanding of the molecular basis of EGFR binding affinities, we docked each EGFR ligand to the putative active state extracellular domain dimer and 25.0 ns molecular dynamics simulations were performed. MM-PBSA/GBSA are efficient computational approaches to approximate free energies of protein-protein interactions and decompose the free energy at the amino acid level. We applied these methods to the last 6.0 ns of each ligand-receptor simulation. MM-PBSA calculations were able to successfully rank all seven of the EGFR ligands based on the two affinity classes: EGF>HB-EGF>TGF-α>BTC>EPR>EPG>AR. Results from energy decomposition identified several interactions that are common among binding ligands. These findings reveal that while several residues are conserved among the EGFR ligand family, no single set of residues determines the affinity class. Instead we found heterogeneous sets of interactions that were driven primarily by electrostatic and Van der Waals forces. These results not only illustrate the complexity of EGFR dynamics but also pave the way for structure-based design of therapeutics targeting EGF ligands or the receptor itself.

Genetic disorders can arise from single base substitutions in a single gene. A single base substitution for wild type guanine in the twelfth codon of KRAS2 mRNA occurs frequently to initiate lung, pancreatic, and colon cancer. We have observed single base mismatch specificity in radioimaging of mutant KRAS2 mRNA in tumors in mice by in vivo hybridization with radiolabeled peptide nucleic acid (PNA) dodecamers. We hypothesized that multi-mutant specificity could be achieved with a PNA dodecamer incorporating hypoxanthine, which can form Watson-Crick basepairs with adenine, cytosine, thymine, and uracil. Using molecular dynamics simulations and free energy calculations, we show that hypoxanthine substitutions in PNAs are tolerated in KRAS2 RNA-PNA duplexes where wild type guanine is replaced by mutant uracil or adenine in RNA. To validate our predictions, we synthesized PNA dodecamers with hypoxanthine, and then measured the thermal stability of RNA-PNA duplexes. Circular dichroism thermal melting results showed that hypoxanthine-containing PNAs are more stable in duplexes where hypoxanthine-adenine and hypoxanthine-uracil base pairs are formed than single mismatch duplexes or duplexes containing hypoxanthine-guanine opposition.

The substitution of natural, bio-based and/or biodegradable polymers for those of petrochemical origin in consumer formulations has become an active area of research and development as the sourcing and destiny...

Molecular dynamics simulations were used to explore two crosslinking mechanisms for in situ formation of polycyanurate thermoset models. Model validation was achieved by comparison of simulated thermophysical properties to experimental values from previous works. Predicted density and coefficient of thermal expansion for the polycyanurate systems agree well with the literature. Glass transition temperature (T g) and water uptake simulations were also consistent with the literature after applying a correction constant. Radial distribution function analysis showed that the absorbed water molecules tended to aggregate in void spaces, specifically around the phenol catalyst, unreacted chain ends, and triazine crosslink moieties. The localization of water around these moieties, as opposed to the canonical bridgehead, suggests an alternative mechanism previously not observed via experimental methods.

Class I aminoacyl-tRNA synthetases (aaRSs) use a Rossmann-fold domain to catalyze the synthesis of aminoacyl-tRNAs required for decoding genetic information. While the Rossmann-fold domain is conserved in evolution, the acceptor stem near the aminoacylation site varies among tRNA substrates, raising the question of how the conserved protein fold adapts to RNA sequence variations. Of interest is the existence of an unpaired C-A mismatch at the 1-72 position unique to bacterial initiator tRNA fMet and absent from elongator tRNAs. Here we show that the class I methionyl-tRNA synthetase (MetRS) of Escherichia coli and its close structural homolog cysteinyl-tRNA synthetase (CysRS) display distinct patterns of recognition of the 1-72 base pair. While the structural homology of the two enzymes in the Rossmann-fold domain is manifested in a common burst feature of aminoacylation kinetics, CysRS discriminates against unpaired 1-72, whereas MetRS lacks such discrimination. A structurebased alignment of the Rossmann fold identifies the insertion of an a-helical motif, specific to CysRS but absent from MetRS, which docks on 1-72 and may discriminate against mismatches. Indeed, substitutions of the CysRS helical motif abolish the discrimination against unpaired 1-72. Additional structural alignments reveal that with the exception of MetRS, class I tRNA synthetases contain a structural motif that docks on 1-72. This work demonstrates that by flexible insertion of a structural motif to dock on 1-72, the catalytic domain of class I tRNA synthetases can acquire structural plasticity to adapt to changes at the end of the tRNA acceptor stem.