Pup-Click-A New Chemoenzymatic Method for the Generation of Singly Pupylated Targets.

Conjugation of the prokaryotic ubiquitin-like protein (Pup) to cellular proteins tags these proteins for degradation by a proteasome in actinobacteria. To study the Pup-proteasome system in biochemical assays, Pup-tagged (i.e.

Proteasome accessory factor A (PafA) transferase activity makes sense in the light of its homology with glutamine synthetase.

The Pup-proteasome system (PPS) is a prokaryotic tagging and degradation system analogous in function to the ubiquitin-proteasome system (UPS). Like ubiquitin, Pup is conjugated to proteins, tagging them for proteasomal degradation. However, in the PPS, a single Pup-ligase, PafA, conjugates Pup to a wide variety of proteins.

How to control an intracellular proteolytic system: Coordinated regulatory switches in the mycobacterial Pup-proteasome system.

Intracellular proteolysis is critical for the proper functioning of all cells, owing to its involvement in a wide range of processes. Because of the destructive nature of protein degradation, intracellular proteolysis is restricted by control mechanisms at almost every step of the proteolytic process. Understanding the coordination of such mechanisms is a challenging task, especially in systems as complex as the eukaryotic ubiquitin-proteasome system (UPS).

An Extended Loop of the Pup Ligase, PafA, Mediates Interaction with Protein Targets.

Pupylation, the bacterial equivalent of ubiquitylation, involves the conjugation of a prokaryotic ubiquitin-like protein (Pup) to protein targets. In contrast to the ubiquitin system, where many ubiquitin ligases exist, a single bacterial ligase, PafA, catalyzes the conjugation of Pup to a wide array of protein targets. As mediators of target recognition by PafA have not been identified, it would appear that PafA alone determines pupylation target selection.

A kinetic model for the prevalence of mono- over poly-pupylation.

Bacteria belonging to the phyla Actinobacteria and Nitrospira possess proteasome cores homologous to the eukaryotic 20S proteasome particle. In these bacteria, the cytoplasmic signal for proteasomal degradation is a small protein termed Pup (prokaryotic ubiquitin-like protein). PafA, the only known Pup ligase, conjugates Pup to lysine side chains of target proteins.