Justin Du Bois

Capturing fleeting intermediates in a catalytic C–H amination reaction cycle

We have applied an ambient ionization technique, desorption electrospray ionization MS, to identify transient reactive species of an archetypal C–H amination reaction catalyzed by a dirhodium tetracarboxylate complex. Using this analytical method, we have detected previously proposed short-lived reaction intermediates, including two nitrenoid complexes that differ in oxidation state. Our findings suggest that an Rh-nitrene oxidant can react with hydrocarbon substrates through a hydrogen atom abstraction pathway and raise the intriguing possibility that two catalytic C–H amination pathways may be operative in a typical bulk solution reaction. As highlighted by these results, desorption electrospray ionization MS should have broad applicability for the mechanistic study of catalytic processes.

Visualizing Dermal Permeation of Sodium Channel Modulators by Mass Spectrometric Imaging

Determining permeability of a given compound through human skin is a principal challenge owing to the highly complex nature of dermal tissue. We describe the application of an ambient mass spectrometry imaging method for visualizing skin penetration of sodium channel modulators, including novel synthetic analogs of natural neurotoxic alkaloids, topically applied ex vivo to human skin. Our simple and label-free approach enables successful mapping of the transverse and lateral diffusion of small molecules having different physicochemical properties without the need for extensive sample preparation.

The paralytic shellfish toxin, saxitoxin, enters the cytoplasm and induces apoptosis of oyster immune cells through a caspase-dependent pathway

Exposure of the toxin-producing dinoflagellate Alexandrium catenella (A. catenella) was previously demonstrated to cause apoptosis of hemocytes in the oyster species Crassostrea gigas. In this work, a coumarin-labeled saxitoxin appeared to spread throughout the cytoplasm of the hemocytes. PSTs, including saxitoxin, were also shown to be directly responsible for inducing apoptosis in hemocytes, a process dependent on caspase activation and independent of reactive oxygen species (ROS) production. A series of in vitro labelling and microscopy experiments revealed that STX and analogs there of induced nuclear condensation, phosphatidylserine exposure, membrane permeability, and DNA fragmentation of hemocytes. Unlike in vertebrates, gonyautoxin-5 (GTX5), which is present in high concentrations in A. catenella, was found to be more toxic than saxitoxin (STX) to oyster immune cells. Altogether, results show that PSTs produced by toxic dinoflagellates enter the cytoplasm and induce apoptosis of oyster immune cells through a caspase-dependent pathway. Because of the central role of hemocytes in mollusc immune defense, PST-induced death of hemocytes could negatively affect resistance of bivalve molluscs to microbial infection.

An expedient synthesis of maraviroc (UK-427,857) via C–H functionalization

A new, concise synthesis of the CCR-5 receptor antagonist maraviroc (UK-427,857) from 3-phenyl-1-propanol has been completed in four steps featuring a site-selective C-H functionalization.

Towards Identifying the Structural Basis For Inhibition By a Newly Discovered Class of CLC Chloride-Channel Inhibitors

CLC chloride channels and transporters play diverse physiological roles. Small-molecule inhibitors of the CLCs would be powerful tools to understand and potentially treat a variety of CLC-related human diseases. In addition to their potential therapeutic benefits, inhibitors can be used to trap channels or transporters in particular conformational states, potentially advancing our understanding of the structural basis for CLC activity. Despite their usefulness, specific small molecule inhibitors for CLC proteins are scarce. To address this shortfall, we have embarked on a search for novel inhibitors of the CLC proteins. We have discovered a panel of small molecules active against both the human ClC-K and the prokaryotic ClC-ec1 homologs. Because ClC-ec1 is amenable to structural studies by X-ray crystallography, co-crystallization experiments with these novel inhibitors are in progress. As an alternative method to identifying the inhibitor-binding site, we have synthesized photo-reactive diazirine derivatives of the inhibitors and shown that these photo-affinity reagents specifically inhibit ClC-ec1. Experiments to identify the binding site using protease digestion coupled with mass spectrometry are currently underway. Once located, protein/inhibitor interactions gleaned from the labeling of ClC-ec1 could allow us to rationally design more potent inhibitors of mammalian transporters and channels.