No abstract

TRIM21 ('tripartite motif-containing protein 21', Ro52) is a ubiquitously expressed cytosolic Fc receptor, which has a potent role in protective immunity against nonenveloped viruses. TRIM21 mediates intracellular neutralisation of antibody-coated viruses, a process called ADIN (antibody-dependent intracellular neutralisation). Our results reveal a similar mechanism to fight bacterial infections. TRIM21 is recruited to the intracellular pathogen Salmonella enterica in epithelial cells early in infection. TRIM21 does not bind directly to S. enterica, but to antibodies opsonising it. Most importantly, bacterial restriction is dependent on TRIM21 as well as on the opsonisation state of the bacteria. Finally, Salmonella and TRIM21 colocalise with the autophagosomal marker LC3, and intracellular defence is enhanced in starved cells suggesting an involvement of the autophagocytic pathway. Our data extend the protective role of TRIM21 from viruses to bacteria and thereby strengthening the general role of ADIN in cellular immunity.

Synthetic biology is the attempt to apply the concepts of engineering to biological systems with the aim to create organisms with new emergent properties. These organisms might have desirable novel biosynthetic capabilities, act as biosensors or help us to understand the intricacies of living systems. This approach has the potential to assist the discovery and production of pharmaceutical compounds at various stages. New sources of bioactive compounds can be created in the form of genetically encoded small molecule libraries. The recombination of individual parts has been employed to design proteins that act as biosensors, which could be used to identify and quantify molecules of interest. New biosynthetic pathways may be designed by stitching together enzymes with desired activities, and genetic code expansion can be used to introduce new functionalities into peptides and proteins to increase their chemical scope and biological stability. This review aims to give an insight into recently developed individual components and modules that might serve as parts in a synthetic biology approach to pharmaceutical biotechnology.

Lysine acylation is a ubiquitous protein modification that controls various aspects of protein function, such as the activity, localization, and stability of enzymes. Mass spectrometric identification of lysine acylations has witnessed tremendous improvements in sensitivity over the last decade, facilitating the discovery of thousands of lysine acylation sites in proteins involved in all essential cellular functions across organisms of all domains of life. However, the vast majority of currently known acylation sites are of unknown function. Semi‐synthetic methods for installing lysine derivatives are ideally suited for in vitro experiments, while genetic code expansion (GCE) allows the installation and study of such lysine modifications, especially their dynamic properties, in vivo. An overview of the current state of the art is provided, and its potential is illustrated with case studies from recent literature. These include the application of engineered enzymes and GCE to install lysine modifications or photoactivatable crosslinker amino acids. Their use in the context of central metabolism, bacterial and viral pathogenicity, the cytoskeleton and chromatin dynamics, is investigated.

The genetic incorporation of unnatural amino acids (UAAs) into proteins by amber suppression technology provides unique avenues to study protein structure, function and interactions both in vitro and in living cells and organisms. This approach has been particularly useful for studying mechanisms of epigenetic chromatin regulation, since these extensively involve dynamic changes in structure, complex formation and posttranslational modifications that are difficult to access by traditional approaches. Here, we review recent achievements in this field, emphasizing UAAs that help to unravel protein-protein interactions in cells by photo-crosslinking or that allow the biosynthesis of proteins with defined posttranslational modifications for studying their function and turnover in vitro and in cells.