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Fast and selective labeling of N-terminal cysteines at neutral pH via thiazolidino boronate formation.

ABSTRACT: Facile labeling of proteins of interest is highly desirable in proteomic research as well as in the development of protein therapeutics. Herein we report a novel method that allows for fast and selective labeling of proteins with an N-terminal cysteine. Although N-terminal cysteines are well known to conjugate with aldehydes to give thiazolidines, the reaction requires acidic conditions and suffers from slow kinetics. We show that benzaldehyde with an ortho-boronic acid substituent readily reacts with N-terminal cysteines at neutral pH, giving rate constants on the order of 10³ M^-1 s^-1. The product features a thiazolidono boronate (TzB) structure and exhibits improved stability due to formation of the B-N dative bond. While stable at neutral pH, the TzB complex dissociates upon mild acidification. These characteristics make the TzB conjugation chemistry potentially useful for the development of drug-protein conjugates that release the small molecule drug in acidic endosomes.

SUBMITTER: Bandyopadhyay A

PROVIDER: S-EPMC5201210 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Fast alpha nucleophiles: structures that undergo rapid hydrazone/oxime formation at neutral pH.

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Fast Diazaborine Formation of Semicarbazide Enables Facile Labeling of Bacterial Pathogens.

Project description:Bioorthogonal conjugation chemistry has enabled the development of tools for the interrogation of complex biological systems. Although a number of bioorthogonal reactions have been documented in literature, they are less ideal for one or several reasons including slow kinetics, low stability of the conjugated product, requirement of toxic catalysts, and side reactions with unintended biomolecules. Herein we report a fast (>103 M-1 s-1) and bioorthogonal conjugation reaction that joins semicarbazide to an aryl ketone or aldehyde with an ortho-boronic acid substituent. The boronic acid moiety greatly accelerates the initial formation of a semicarbazone conjugate, which rearranges into a stable diazaborine. The diazaborine formation can be performed in blood serum or cell lysates with minimal interference from biomolecules. We further demonstrate that application of this conjugation chemistry enables facile labeling of bacteria. A synthetic amino acid D-AB3, which presents a 2-acetylphenylboronic acid moiety as its side chain, was found to incorporate into several bacterial species through cell wall remodeling, with particularly high efficiency for Escherichia coli. Subsequent D-AB3 conjugation to a fluorophore-labeled semicarbazide allows robust detection of this bacterial pathogen in blood serum.

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pH Responsive Antibacterial Hydrogel Utilizing Catechol-Boronate Complexation Chemistry.

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