We report the measurement and analysis
of sulfonium−π,
thioether−π, and ammonium−π interactions
in a β-hairpin peptide model system, coupled with computational
investigation and PDB analysis. These studies indicated that the sulfonium−π
interaction is the strongest and that polarizability contributes to
the stronger interaction with sulfonium relative to ammonium. Computational
studies demonstrate that differences in solvation of the trimethylsulfonium
versus the trimethylammonium group also contribute to the stronger
sulfonium−π interaction. In comparing sulfonium−π
versus sulfur−π interactions in proteins, analysis of
SAM- and SAH-bound enzymes in the PDB suggests that aromatic residues
are enriched in close proximity to the sulfur of both SAM and SAH,
but the populations of aromatic interactions of the two cofactors
are not significantly different, with the exception of the Me−π
interactions in SAM, which are the most prevalent interaction in SAM
but are not possible for SAH. This suggests that the weaker interaction
energies due to loss of the cation−π interaction in going
from SAM to SAH may contribute to turnover of the cofactor.
mRNA display of macrocyclic peptides has proven itself to be a powerful technique to discover high-affinity ligands for a protein target. However, only a limited number of cyclization chemistries are known to be compatible with mRNA display. Tyrosinase is a copper-dependent oxidase that oxidizes tyrosine phenol to an electrophilic o-quinone, which is readily attacked by cysteine thiol. Here we show that peptides containing tyrosine and cysteine are rapidly cyclized upon tyrosinase treatment. Characterization of the cyclization reveals it to be widely applicable to multiple macrocycle sizes and scaffolds. We combine tyrosinase-mediated cyclization with mRNA display to discover new macrocyclic ligands targeting melanoma-associated antigen A4 (MAGE-A4). These macrocycles potently inhibit the MAGE-A4 binding axis with nanomolar IC 50 values. Importantly, macrocyclic ligands show clear advantage over noncyclized analogues with ∼40-fold or greater decrease in IC 50 values.
Background: Comparability of cholesterol measurement is clinically required and external quality assurance (EQA) programmes are important to verify the trueness of routine methods. Methods: We developed a gas chromatography-isotope dilution mass spectrometry (GC-IDMS) total cholesterol assay to investigate the cause of a suspected matrix-related negative bias with the Beckman Coulter enzymatic method discovered in an EQA programme. The GC-IDMS method was calibrated with certified reference material and verified against a secondary reference method. Bias between the GC-IDMS and Beckman Coulter methods was estimated according to Clinical and Laboratory Standards Institute (CLSI) protocol EP9-A2 with 40 clinical samples. Results: At clinically important decision levels, no significant bias was demonstrated on patients' samples (all results within a +3% limit). A matrix effect confined to the EQA material that affected the Beckman Coulter total cholesterol method was confirmed. Conclusions: The GC-IDMS method is suitable as a higher order total cholesterol method in a routine clinical laboratory. Matrix effects defeat the objectives of EQA schemes by preventing the verification of trueness. Given the importance of obtaining a true cholesterol result without systematic error, we recommend that EQA material without matrix effects should be used.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.