The Bacterial Proteasome at the Core of Diverse Degradation Pathways

Proteasomal protein degradation exists in mycobacteria and other actinobacteria, and expands their repertoire of compartmentalizing protein degradation pathways beyond the usual bacterial types. A product of horizontal gene transfer, bacterial proteasomes have evolved to support the organism's survival under challenging environmental conditions like nutrient starvation and physical or chemical stresses. Like the eukaryotic 20S proteasome, the bacterial core particle is gated and must associate with a regulator complex to form a fully active protease capable of recruiting and internalizing substrate proteins. By association with diverse regulator complexes that employ different recruitment strategies, the bacterial 20S core particle is able to act in different cellular degradation pathways. In association with the mycobacterial proteasomal ATPase Mpa, the proteasome degrades substrates post-translationally modified with prokaryotic, ubiquitin-like protein Pup in a process called pupylation. Upon interaction with the ATP-independent bacterial proteasome activator Bpa, poorly structured substrates are recruited for proteasomal degradation. A potential third degradation route might employ a Cdc48-like protein of actinobacteria (Cpa), for which interaction with the 20S core was recently demonstrated but no degradation substrates have been identified yet. The alternative interaction partners and wide range of substrate proteins suggest that the bacterial proteasome is a modular, functionally flexible and conditionally regulated degradation machine in bacteria that encounter rapidly changing and challenging conditions

A distinct structural region of the prokaryotic ubiquitin-like protein (Pup) is recognized by the N-terminal domain of the proteasomal ATPase Mpa

AbstractThe mycobacterial ubiquitin-like protein Pup is coupled to proteins, thereby rendering them as substrates for proteasome-mediated degradation. The Pup-tagged proteins are recruited by the proteasomal ATPase Mpa (also called ARC). Using a combination of biochemical and NMR methods, we characterize the structural determinants of Pup and its interaction with Mpa, demonstrating that Pup adopts a range of extended conformations with a short helical stretch in its C-terminal portion. We show that the N-terminal coiled-coil domain of Mpa makes extensive contacts along the central region of Pup leaving its N-terminus unconstrained and available for other functional interactions.Structured summaryMINT-7262427: pup (uniprotkb:B6DAC1) binds (MI:0407) to mpa (uniprotkb:Q0G9Y7) by pull down (MI:0096) MINT-7262440: mpa (uniprotkb:Q0G9Y7) and pup (uniprotkb:B6DAC1) bind (MI:0407) by isothermal titration calorimetry (MI:0065

Pupylation-dependent and -independent proteasomal degradation in mycobacteria

Bacteria make use of compartmentalizing protease complexes, similar in architecture but not homologous to the eukaryotic proteasome, for the selective and processive removal of proteins. Mycobacteria as members of the actinobacteria harbor proteasomes in addition to the canonical bacterial degradation complexes. Mycobacterial proteasomal degradation, although not essential during normal growth, becomes critical for survival under particular environmental conditions, like, for example, during persistence of the pathogenic Mycobacterium tuberculosis in host macrophages or of environmental mycobacteria under starvation. Recruitment of protein substrates for proteasomal degradation is usually mediated by pupylation, the post-translational modification of lysine side chains with the prokaryotic ubiquitin-like protein Pup. This substrate recruitment strategy is functionally reminiscent of ubiquitination in eukaryotes, but is the result of convergent evolution, relying on chemically and structurally distinct enzymes. Pupylated substrates are recognized by the ATP-dependent proteasomal regulator Mpa that associates with the 20S proteasome core. A pupylation-independent proteasome degradation pathway has recently been discovered that is mediated by the ATP-independent bacterial proteasome activator Bpa (also referred to as PafE), and that appears to play a role under stress conditions. In this review, mechanistic principles of bacterial proteasomal degradation are discussed and compared with functionally related elements of the eukaryotic ubiquitin-proteasome system. Special attention is given to an understanding on the molecular level based on structural and biochemical analysis. Wherever available, discussion of in vivo studies is included to highlight the biological significance of this unusual bacterial degradation pathway

Pupylation-dependent and -independent proteasomal degradation in mycobacteria

Bacteria make use of compartmentalizing protease complexes, similar in architecture but not homologous to the eukaryotic proteasome, for the selective and processive removal of proteins. Mycobacteria as members of the actinobacteria harbor proteasomes in addition to the canonical bacterial degradation complexes. Mycobacterial proteasomal degradation, although not essential during normal growth, becomes critical for survival under particular environmental conditions, like, for example, during persistence of the pathogenic Mycobacterium tuberculosis in host macrophages or of environmental mycobacteria under starvation. Recruitment of protein substrates for proteasomal degradation is usually mediated by pupylation, the post-translational modification of lysine side chains with the prokaryotic ubiquitin-like protein Pup. This substrate recruitment strategy is functionally reminiscent of ubiquitination in eukaryotes, but is the result of convergent evolution, relying on chemically and structurally distinct enzymes. Pupylated substrates are recognized by the ATP-dependent proteasomal regulator Mpa that associates with the 20S proteasome core. A pupylation-independent proteasome degradation pathway has recently been discovered that is mediated by the ATP-independent bacterial proteasome activator Bpa (also referred to as PafE), and that appears to play a role under stress conditions. In this review, mechanistic principles of bacterial proteasomal degradation are discussed and compared with functionally related elements of the eukaryotic ubiquitin-proteasome system. Special attention is given to an understanding on the molecular level based on structural and biochemical analysis. Wherever available, discussion of in vivo studies is included to highlight the biological significance of this unusual bacterial degradation pathway

Survival in Hostile Conditions: Pupylation and the Proteasome in Actinobacterial Stress Response Pathways

Bacteria employ a multitude of strategies to cope with the challenges they face in their natural surroundings, be it as pathogens, commensals or free-living species in rapidly changing environments like soil. Mycobacteria and other Actinobacteria acquired proteasomal genes and evolved a post-translational, ubiquitin-like modification pathway called pupylation to support their survival under rapidly changing conditions and under stress. The proteasomal 20S core particle (20S CP) interacts with ring-shaped activators like the hexameric ATPase Mpa that recruits pupylated substrates. The proteasomal subunits, Mpa and pupylation enzymes are encoded in the so-called Pup-proteasome system (PPS) gene locus. Genes in this locus become vital for bacteria to survive during periods of stress. In the successful human pathogen Mycobacterium tuberculosis, the 20S CP is essential for survival in host macrophages. Other members of the PPS and proteasomal interactors are crucial for cellular homeostasis, for example during the DNA damage response, iron and copper regulation, and heat shock. The multiple pathways that the proteasome is involved in during different stress responses suggest that the PPS plays a vital role in bacterial protein quality control and adaptation to diverse challenging environments

Conditional displacement operator for traveling fields

We show that the conditional displacement operator $\hat{U}_{CD}=\exp [\hat{b}^{\dagger}\hat{b}(\beta \hat{a}^{\dagger}-\beta ^{\ast}\hat{a})]$ acting upon an arbitrary state of traveling waves can be well approximated by the action of a Kerr medium placed between two beam splitters whose respective second ports are fed by highly excited coherent states. Applications to the generation of nonclassical states and measurement of Wigner function of arbitrary states are also considered

SoLid : Search for Oscillations with Lithium-6 Detector at the SCK-CEN BR2 reactor

Sterile neutrinos have been considered as a possible explanation for the recent reactor and Gallium anomalies arising from reanalysis of reactor flux and calibration data of previous neutrino experiments. A way to test this hypothesis is to look for distortions of the anti-neutrino energy caused by oscillation from active to sterile neutrino at close stand-off (similar to 6-8m) of a compact reactor core. Due to the low rate of anti-neutrino interactions the main challenge in such measurement is to control the high level of gamma rays and neutron background. The SoLid experiment is a proposal to search for active-to-sterile anti-neutrino oscillation at very short baseline of the SCK center dot CEN BR2 research reactor. This experiment uses a novel approach to detect anti-neutrino with a highly segmented detector based on Lithium-6. With the combination of high granularity, high neutron-gamma discrimination using 6LiF:ZnS(Ag) and precise localization of the Inverse Beta Decay products, a better experimental sensitivity can be achieved compared to other state-of-the-art technology. This compact system requires minimum passive shielding allowing for very close stand off to the reactor. The experimental set up of the SoLid experiment and the BR2 reactor will be presented. The new principle of neutrino detection and the detector design with expected performance will be described. The expected sensitivity to new oscillations of the SoLid detector as well as the first measurements made with the 8 kg prototype detector deployed at the BR2 reactor in 2013-2014 will be reported

Thermotolerance Requires Refolding of Aggregated Proteins by Substrate Translocation through the Central Pore of ClpB

AbstractCell survival under severe thermal stress requires the activity of the ClpB (Hsp104) AAA+ chaperone that solubilizes and reactivates aggregated proteins in concert with the DnaK (Hsp70) chaperone system. How protein disaggregation is achieved and whether survival is solely dependent on ClpB-mediated elimination of aggregates or also on reactivation of aggregated proteins has been unclear. We engineered a ClpB variant, BAP, which associates with the ClpP peptidase and thereby is converted into a degrading disaggregase. BAP translocates substrates through its central pore directly into ClpP for degradation. ClpB-dependent translocation is demonstrated to be an integral part of the disaggregation mechanism. Protein disaggregation by the BAP/ClpP complex remains dependent on DnaK, defining a role for DnaK at early stages of the disaggregation reaction. The activity switch of BAP to a degrading disaggregase does not support thermotolerance development, demonstrating that cell survival during severe thermal stress requires reactivation of aggregated proteins

Strange Quark Matter and Compact Stars

Astrophysicists distinguish between three different types of compact stars. These are white dwarfs, neutron stars, and black holes. The former contain matter in one of the densest forms found in the Universe which, together with the unprecedented progress in observational astronomy, make such stars superb astrophysical laboratories for a broad range of most striking physical phenomena. These range from nuclear processes on the stellar surface to processes in electron degenerate matter at subnuclear densities to boson condensates and the existence of new states of baryonic matter--like color superconducting quark matter--at supernuclear densities. More than that, according to the strange matter hypothesis strange quark matter could be more stable than nuclear matter, in which case neutron stars should be largely composed of pure quark matter possibly enveloped in thin nuclear crusts. Another remarkable implication of the hypothesis is the possible existence of a new class of white dwarfs. This article aims at giving an overview of all these striking physical possibilities, with an emphasis on the astrophysical phenomenology of strange quark matter. Possible observational signatures associated with the theoretically proposed states of matter inside compact stars are discussed as well. They will provide most valuable information about the phase diagram of superdense nuclear matter at high baryon number density but low temperature, which is not accessible to relativistic heavy ion collision experiments.Comment: 58 figures, to appear in "Progress in Particle and Nuclear Physics"; References added for sections 1,2,3,5; Equation (116) corrected; Figs. 1 and 58 update