Mechanism of autocatalytic activation during proteasome assembly.

Many large molecular machines are too elaborate to assemble spontaneously and are built through ordered pathways orchestrated by dedicated chaperones. During assembly of the core particle (CP) of the proteasome, where protein degradation occurs, its six active sites are simultaneously activated via cleavage of N-terminal propeptides. Such activation is autocatalytic and coupled to fusion of two half-CP intermediates, which protects cells by preventing activation until enclosure of the active sites within the CP interior.

Rational design of proteasome inhibitors based on the structure of the endogenous inhibitor PI31/Fub1.

Proteasome inhibitors are widely used anticancer drugs. The three clinically approved agents are modified small peptides that preferentially target one of the proteasome's three active sites (β5) at physiologic concentrations. In addition to these drugs, there is also an endogenous proteasome inhibitor, PI31/Fub1, that enters the proteasome's interior to simultaneously yet specifically inhibit all three active sites.

Experimental and Computational Observations of Immunogenic Cobalt Porphyrin Lipid Bilayers: Nanodomain-Enhanced Antigen Association.

Cobalt porphyrin phospholipid (CoPoP) can incorporate within bilayers to enable non-covalent surface-display of antigens on liposomes by mixing with proteins bearing a polyhistidine tag (his-tag); however, the mechanisms for how this occurs are poorly understood. These were investigated using the his-tagged model antigen Pfs25, a protein antigen candidate for malaria transmission-blocking vaccines. Pfs25 was found to associate with the small molecule aquocobalamin, a form of vitamin B12 and a cobalt-containing corrin macrocycle, but without particle formation, enabling comparative assessment.

Structural consequences of the interaction of RbgA with a 50S ribosomal subunit assembly intermediate.

Bacteria harbor a number GTPases that function in the assembly of the ribosome and are essential for growth. RbgA is one of these GTPases and is required for the assembly of the 50S subunit in most bacteria. Homologs of this protein are also implicated in the assembly of the large subunit of the mitochondrial and eukaryotic ribosome.

Streptomyces IHF uses multiple interfaces to bind DNA.

Background: Nucleoid associated proteins (NAPs) are essential for chromosome condensation in bacterial cells. Despite being a diverse group, NAPs share two common traits: they are small, oligomeric proteins and their oligomeric state is critical for DNA condensation. Streptomyces coelicolor IHF (sIHF) is an actinobacterial-specific nucleoid-associated protein that despite its name, shares neither sequence nor structural homology with the well-characterized Escherichia coli IHF.

Role of Era in assembly and homeostasis of the ribosomal small subunit.

Assembly factors provide speed and directionality to the maturation process of the 30S subunit in bacteria. To gain a more precise understanding of how these proteins mediate 30S maturation, it is important to expand on studies of 30S assembly intermediates purified from bacterial strains lacking particular maturation factors. To reveal the role of the essential protein Era in the assembly of the 30S ribosomal subunit, we analyzed assembly intermediates that accumulated in Era-depleted Escherichia coli cells using quantitative mass spectrometry, high resolution cryo-electron microscopy and in-cell footprinting.

Highly-Soluble Cyanine J-aggregates Entrapped by Liposomes for Optical Imaging around 930 nm.

Near infrared (NIR) dyes are useful for optical imaging. Liposomes have been used extensively for delivery of diverse cargos, including hydrophilic cargos which are passively loaded in the aqueous core. However, most currently available NIR dyes are only slightly soluble in water, making passive entrapment in liposomes challenging for achieving high optical contrast.

A malaria vaccine adjuvant based on recombinant antigen binding to liposomes.

Pfs25 is a malaria transmission-blocking vaccine antigen candidate, but its apparently limited immunogenicity in humans has hindered clinical development. Here, we show that recombinant, polyhistidine-tagged (his-tagged) Pfs25 can be mixed at the time of immunization with pre-formed liposomes containing cobalt porphyrin-phospholipid, resulting in spontaneous nanoliposome antigen particleization (SNAP). Antigens are stably presented in uniformly orientated display via his-tag insertion in the cobalt porphyrin-phospholipid bilayer, without covalent modification or disruption of antigen conformation.

The cryo-EM structure of YjeQ bound to the 30S subunit suggests a fidelity checkpoint function for this protein in ribosome assembly.

Recent work suggests that bacterial YjeQ (RsgA) participates in the late stages of assembly of the 30S subunit and aids the assembly of the decoding center but also binds the mature 30S subunit with high affinity. To determine the function and mechanisms of YjeQ in the context of the mature subunit, we determined the cryo-EM structure of the fully assembled 30S subunit in complex with YjeQ at 5.8-Å resolution.

Final touches and quality control on the assembly of the eukaryotic ribosome.

One of the most fundamental processes of life is protein synthesis by the ribosome. Although much is known about the function and structure of this macromolecular complex, our understanding on its assembly is still vague. In this issue of , Malyutin (2017) provide a detailed picture of one of the latest assembly stages of the yeast 60S ribosomal subunit.

The impact of recent improvements in cryo-electron microscopy technology on the understanding of bacterial ribosome assembly.

Cryo-electron microscopy (cryo-EM) had played a central role in the study of ribosome structure and the process of translation in bacteria since the development of this technique in the mid 1980s. Until recently cryo-EM structures were limited to ∼10 Å in the best cases. However, the recent advent of direct electron detectors has greatly improved the resolution of cryo-EM structures to the point where atomic resolution is now achievable.

Sphingomyelin Liposomes Containing Porphyrin-phospholipid for Irinotecan Chemophototherapy.

Porphyrin-phospholipid (PoP) liposomes can entrap anti-cancer agents and release them in response to near infrared (NIR) light. Doxorubicin, when remotely loaded via an ammonium sulfate gradient at a high drug-to-lipid ratio, formed elongated crystals that altered liposome morphology and could not be loaded into liposomes with higher PoP content. On the other hand, irinotecan could also be remotely loaded but did not form large crystals and did not induce liposome elongation.

YphC and YsxC GTPases assist the maturation of the central protuberance, GTPase associated region and functional core of the 50S ribosomal subunit.

YphC and YsxC are GTPases in Bacillus subtilis that facilitate the assembly of the 50S ribosomal subunit, however their roles in this process are still uncharacterized. To explore their function, we used strains in which the only copy of the yphC or ysxC genes were under the control of an inducible promoter. Under depletion conditions, they accumulated incomplete ribosomal subunits that we named 45SYphC and 44.

Lack of Adipocyte AMPK Exacerbates Insulin Resistance and Hepatic Steatosis through Brown and Beige Adipose Tissue Function.

Brown (BAT) and white (WAT) adipose tissues play distinct roles in maintaining whole-body energy homeostasis, and their dysfunction can contribute to non-alcoholic fatty liver disease (NAFLD) and type 2 diabetes. The AMP-activated protein kinase (AMPK) is a cellular energy sensor, but its role in regulating BAT and WAT metabolism is unclear. We generated an inducible model for deletion of the two AMPK β subunits in adipocytes (iβ1β2AKO) and found that iβ1β2AKO mice were cold intolerant and resistant to β-adrenergic activation of BAT and beiging of WAT.

Porphyrin-phospholipid liposomes with tunable leakiness.

Drug bioavailability is a key consideration for drug delivery systems. When loaded with doxorubicin, liposomes containing 5 molar % porphyrin-phospholipid (HPPH liposomes) exhibited in vitro and in vivo serum stability that could be fine-tuned by varying the drug-to-lipid ratio. A higher drug loading ratio destabilized the liposomes, in contrast to standard liposomes which displayed an opposite and less pronounced trend.

Doxorubicin encapsulated in stealth liposomes conferred with light-triggered drug release.

Stealth liposomes can be used to extend the blood circulation time of encapsulated therapeutics. Inclusion of 2 molar % porphyrin-phospholipid (PoP) imparted optimal near infrared (NIR) light-triggered release of doxorubicin (Dox) from conventional sterically stabilized stealth liposomes. The type and amount of PoP affected drug loading, serum stability and drug release induced by NIR light.

The C-terminal helix in the YjeQ zinc-finger domain catalyzes the release of RbfA during 30S ribosome subunit assembly.

YjeQ (also called RsgA) and RbfA proteins in Escherichia coli bind to immature 30S ribosome subunits at late stages of assembly to assist folding of the decoding center. A key step for the subunit to enter the pool of actively translating ribosomes is the release of these factors. YjeQ promotes dissociation of RbfA during the final stages of maturation; however, the mechanism implementing this functional interplay has not been elucidated.