John C. Jewett

Water‐Soluble Triazabutadienes that Release Diazonium Species upon Protonation under Physiologically Relevant Conditions

Triazabutadienes are an understudied structural motif that have remarkable reactivity once rendered water-soluble. It is shown that these molecules readily release diazonium species in a pH-dependent manner in a series of buffer solutions with pH ranges similar to those found in cells. Upon further development, we expect that this process will be well suited to cargo-release strategies and organelle-specific bioconjugation reactions. These compounds offer one of the mildest ways of generating diazonium species in aqueous solutions.

Light-Activated Triazabutadienes for the Modification of a Viral Surface.

Chemical crosslinking is a versatile tool for the examination of biochemical interactions, in particular host–pathogen interactions. We report the critical first step toward the goal of probing these interactions by the synthesis and use of a new heterobifunctional crosslinker containing a triazabutadiene scaffold. The triazabutadiene is stable to protein conjugation and liberates a reactive aryl diazonium species upon irradiation with 350 nm light. We highlight the use of this technology by modifying the surface of several proteins, including the dengue virus envelope protein.

Protecting triazabutadienes to afford acid resistance

Recent work on triazabutadienes has shown that they have the ability to release aryl diazonium ions under exceptionally mild acidic conditions. There are instances that require that this release be prevented or minimized. Accordingly, a base‐labile protection strategy for the triazabutadiene is presented. It affords enhanced synthetic and practical utility of the triazabutadiene. The effects of steric and electronic factors in the rate of removal are discussed, and the triazabutadiene protection is shown to be compatible with the traditional acid‐labile protection strategy used in solid phase peptide synthesis.

A gradient-free method for the purification of infective dengue virus for protein-level investigations.

Dengue virus (DENV) is a mosquito-transmitted flavivirus that infects approximately 100 million people annually. Multi-day protocols for purification of DENV reduce the infective titer due to viral sensitivity to both temperature and pH. Herein we describe a 5-h protocol for the purification of all DENV serotypes, utilizing traditional gradient-free ultracentrifugation followed by selective virion precipitation. This protocol allows for the separation of DENV from contaminating proteins - including intact C6/36 densovirus, for the production of infective virus at high concentration for protein-level analysis.

Coumarin triazabutadienes for fluorescent labeling of proteins

The use of small‐molecule fluorophores to label proteins with minimal perturbation in response to an external stimulus is a powerful tool to probe chemical and biochemical environments. Herein, we describe the use of a coumarin‐modified triazabutadiene that can deliver aryl diazonium ions to fluorescently label proteins by tyrosine‐selective modification. The labeling can be triggered by low‐pH‐induced liberation of the diazonium species, thus making the fluorophore especially useful in labeling biochemical surroundings such as those found within the late endosome. Additionally, we show that a variety of coumarin triazabutadienes might also be prone to releasing their diazonium cargo after irradiation with UV light.