Recent advances of thiol-selective bioconjugation reactions.

Proteins are the most abundant biomolecules within a cell and are involved in all biochemical cellular processes, fulfilling specific functions with unmatched precision. This unique specificity makes proteins an ideal scaffold to generate tools for the exploration of natural systems or for the construction of modern therapeutics. Thus, the chemoselective modification of proteins with functionalities that are not defined by the genetic code has become an indispensable approach for life science research and the development of therapeutics. Amongst site-selective strategies for protein modification, cysteine-selective approaches have long been used for the generation of functional protein conjugates and new reactions continue to emerge, offering solutions for diverse research questions. In this review, we are highlighting new strategies for the chemoselective modification of cysteine residues in peptides, proteins and antibodies with a particular focus on the most recent years. We lay special focus on new reagents for efficient cysteine conjugation that produce stable conjugation products with significant pharmaceutical application.

Tag and release: strategies for the intracellular cleavage of protein conjugates.

Attaching a functional moiety to a protein is required for a wealth of applications, comprising targeted delivery, controlling of enzyme activity, and prodrug-based therapy. Targeting intracellular processes requires the cellular delivery of the protein. While at first, a stable connection between the protein and the modification is desired, once inside the cell, the conjugate might be cleaved again to restore or activate the function of the individual parts. This can be achieved by employing cleavable linkages in conjugates, which are responsive to chemical or enzymatic stimuli inside cells. In this overview we describe strategies, how such entities can be introduced into proteins and how selective intracellular cleavage can be accomplished.

Fluorescent labelling in living cells.

The labelling of proteins with green fluorescent protein enabled the visualization of proteins in living cells for the first time. Since then, much progress has been made in the field. Modern strategies allow the labelling of proteins in live cells through a range of specialized methods with sophisticated chemical probes that show enhanced photophysical properties compared to fluorescent proteins. This review briefly summarizes recent advances in the field of fluorescent chemical protein labelling inside living cells and illustrates key aspects on the requirements and advantages of each given method.