Site-specific protein conjugates incorporating Para-Azido-L-Phenylalanine for cellular and in vivo imaging.

This work features the use of amber suppression-mediated unnatural amino acid (UAA) incorporation into proteins for various imaging purposes. The site-specific incorporation of the UAA, p-azido-L-phenylalanine (pAzF), provides an azide handle that can be used to complete the strain promoted azide-alkyne click cycloaddition (SPAAC) reaction to introduce an imaging modality such as a fluorophore or a positron emission tomography (PET) tracer on the protein of interest (POI). Such methodology can be pursued directly in mammalian cell lines or on proteins expressed in vitro, thereby conferring a homogeneous pool of protein conjugates. A general procedure for UAA incorporation to use with a site-specific protein labeling method is provided allowing for in vitro and in vivo imaging applications based on the representative proteins PTEN and PD-L1. This approach would help elucidate the cellular or in vivo biological activities of the POI.

Unprotected peptide macrocyclization and stapling via a fluorine-thiol displacement reaction.

We report the discovery of a facile peptide macrocyclization and stapling strategy based on a fluorine thiol displacement reaction (FTDR), which renders a class of peptide analogues with enhanced stability, affinity, cellular uptake, and inhibition of cancer cells. This approach enabled selective modification of the orthogonal fluoroacetamide side chains in unprotected peptides in the presence of intrinsic cysteines. The identified benzenedimethanethiol linker greatly promoted the alpha helicity of a variety of peptide substrates, as corroborated by molecular dynamics simulations. The cellular uptake of benzenedimethanethiol stapled peptides appeared to be universally enhanced compared to the classic ring-closing metathesis (RCM) stapled peptides. Pilot mechanism studies suggested that the uptake of FTDR-stapled peptides may involve multiple endocytosis pathways in a distinct pattern in comparison to peptides stapled by RCM. Consistent with the improved cell permeability, the FTDR-stapled lead Axin and p53 peptide analogues demonstrated enhanced inhibition of cancer cells over the RCM-stapled analogues and the unstapled peptides.

Capture and Kill: Selective Eradication of Target Bacteria by a Flexible Bacteria-Imprinted Chip.

This paper reports an antibacterial chip that can selectively capture bacteria and kill them using low-voltage DC electricity. We prepared a bacteria-imprinted, flexible PDMS chip that can separate target bacteria from suspensions with high selectivity. The chip contained integrated electrodes that can kill the captured bacteria within 10 min by applying a low DC voltage. The used chip could be easily regenerated by solution immersion. Meanwhile, the PDMS chip showed good biocompatibility and inhibited adhesion of human blood cells. Our work points to a new strategy to address pathogenic bacterial contamination and infection.