The mitochondrial channel VDAC has a cation-selective open state

The mitochondrial channel VDAC is known to have two major classes of functional states, a large conductance "open" state that is anion selective, and lower conductance substates that are cation selective. The channel can reversibly switch between open and half-open states, with the latter predominant at increasing membrane voltages of either polarity. We report the presence of a new functional state of VDAC, a cation-selective state with conductance approximately equal to that of the canonical open state. This newly described state of VDAC can be reached from either the half-open cation-selective state or from the open anion-selective state. The latter transition implies that a mechanism exists for selectivity gating in VDAC that is separate from partial closure, which may be relevant to the physiological regulation of this channel and mitochondrial outer membrane permeability.     

The structure of the O-polysaccharide of the Pseudoalteromonas rubra ATCC 29570T lipopolysaccharide containing a keto sugar

The structure of the phenol-soluble polysaccharide from Pseudoalteromonas rubra type strain ATCC 29570T has been elucidated using 1H and 13C NMR spectroscopy, including 2D COSY, TOCSY, gNOESY, ROESY, 1H,13C gHMQC and gHMBC experiments. It is concluded that the trisaccharide repeating unit of the polysaccharide has the following structure: [carbohydrate structure: see text] where Sug is 2-acetamido-2,6-dideoxy-D-xylo-hexos-4-ulose, Am is acetimidoyl and Acyl is a malic acid residue, which is O-acetylated in approximately 70% of the units.     

The structure of the glucuronoxylomannan produced by culinary-medicinal yellow brain mushroom (Tremella mesenterica Ritz.:Fr., Heterobasidiomycetes) grown as one cell biomass in submerged culture

The yellow brain mushroom Tremella mesenterica possesses a wide spectrum of medicinal properties, including immunostimulating, protecting against radiation, antidiabetic, anti-inflammatory, hypocholesterolemic, hepatoprotective, and antiallergic effects. A unique feature of T. mesenterica is that most of the above mentioned medicinal properties depend on glucuronoxylomannan (GXM) contained in fruiting bodies or produced in pure culture conditions. We developed a new strain of T. mesenterica CBS 101939, which grows in submerged culture and offers superior yields of one-cell biomass rich in exocellular heteropolysaccharide GXM. The structure of the GXM was analyzed by NMR spectroscopy and chemical methods. The polysaccharide has a defined repeating unit structure, which is O-acetylated at several points: [structure: see text]. These results differ from previously published structure of Tremella extracellular polysaccharides, where mannan backbone was believed to be randomly glycosylated with xylan chains of different length.     

Structure of an acidic O-specific polysaccharide from marine bacterium Shewanella fidelis KMM 3582T containing Nepsilon-[(S)-1-carboxyethyl]-Nalpha-(D-galacturonoyl)-L-lysine

The O-specific polysaccharide was isolated by mild acid degradation of the lipopolysaccharide of the marine bacterium Shewanella fidelis type strain KMM 3582T and studied by sugar analysis along with 1H and 13C NMR spectroscopy including one-dimensional NOE in difference mode and two-dimensional experiments. The polysaccharide was found to consist of linear tetrasaccharide repeating units containing Nepsilon-[(S)-1-carboxyethyl]-Nalpha-(D-galacturonoyl)-L-lysine and having the following structure: [See text.] The amide of D-galacturonic acid with Nepsilon-[(S)-1-carboxyethyl]-L-lysine ('alaninolysine', 2S,8S-AlaLys) was found for the first time in nature as a component of the O-specific polysaccharide of Providencia rustigianii O14 (Carbohydr. Res. 2003, 338, 1009-1016).     

Mycobactin-mediated iron acquisition within macrophages

Restricting the availability of iron is an important strategy for defense against bacterial infection. Mycobacterium tuberculosis survives within the phagosomes of macrophages; consequently, iron acquisition is particularly difficult for M. tuberculosis, because the phagosomal membrane is an additional barrier for its iron access. However, little is known about the iron transport and acquisition pathways adapted by this microbe in vivo. Extracellular iron sources are usually mobilized by hydrophilic siderophores. Here, we describe direct evidence that mycobactins, the lipophilic siderophores of mycobacteria, efficiently extract intracellular macrophage iron. The metal-free siderophore is diffusely associated with the macrophage membrane, ready for iron chelation. Notably, the mycobactin-metal complex accumulates with high selectivity in macrophage lipid droplets, intracellular domains for lipid storage and sorting. In our experiments, these mycobactin-targeted lipid droplets were found in direct contact with phagosomes, poised for iron delivery. The existence of this previously undescribed iron acquisition pathway indicates that mycobacteria have taken advantage of endogenous macrophage mechanisms for iron mobilization and lipid sorting for iron acquisition during infection. The pathway could represent a new target for the control of mycobacterial infection.     

Lipopolysaccharides of anaerobic beer spoilage bacteria of the genus Pectinatus--lipopolysaccharides of a Gram-positive genus

Bacteria of the genus Pectinatus emerged during the seventies as contaminants and spoilage organisms in packaged beer. This genus comprises two species, Pectinatus cerevisiiphilus and Pectinatus frisingensis; both are strict anaerobes. On the basis of genomic properties the genus is placed among low GC Gram-positive bacteria (phylum Firmicutes, class Clostridia, order Clostridiales, family Acidaminococcaceae). Despite this assignment, Pectinatus bacteria possess an outer membrane and lipopolysaccharide (LPS) typical of Gram-negative bacteria. The present review compiles the structural and compositional studies performed on Pectinatus LPS. These lipopolysaccharides exhibit extensive heterogeneity, i.e. several macromolecularly and structurally distinct LPS molecules are produced by each strain. Whereas heterogeneity is a common property in lipopolysaccharides, Pectinatus LPS have been shown to contain exceptional carbohydrate structures, consisting of a fairly conserved core region that carries a large non-repetitive saccharide that probably replaces the O-specific chain. Such structures represent a novel architectural principle of the LPS molecule.     

A Model of Inositol Compartmentation in Astrocytes Based Upon Efflux Kinetics and Slow Inositol Depletion after Uptake Inhibition

Intracellular compartmentation of inositol was demonstrated in primary cultures of mouse astrocytes, incubated in isotonic medium, by determination of efflux kinetics after loading with [³H]inositol. Three kinetically different compartments were delineated. The largest and most slowly exchanging compartment had a halflife of 9 hr. This slow release leads to retention of a sizeable amount of pre-accumulated inositol in the tissue 24 hr after the onset of uptake inhibition, as confirmed by the observation that the inositol uptake inhibitor fucose caused a larger inhibition of unidirectional inositol uptake than of inositol pool size, measured as accumulated [³H]inositol after 24 hr of combined exposure to the inhibitor and the labeled isotope. Based upon the present observations and literature data, it is suggested that the large, slowly exchanging compartment is largely membrane-associated and participating in signaling via the phosphatidylinositide second messenger system, whereas inositol functioning as an osmolyte is distributed in the cytosol and located in one or both of the compartments showing a faster release.     

Interaction of the signaling state analog and the apoprotein form of the orange carotenoid protein with the fluorescence recovery protein

Photoprotection in cyanobacteria relies on the interplay between the orange carotenoid protein (OCP) and the fluorescence recovery protein (FRP) in a process termed non-photochemical quenching, NPQ. Illumination with blue-green light converts OCP from the basic orange state (OCP^O) into the red-shifted, active state (OCP^R) that quenches phycobilisome (PBs) fluorescence to avoid excessive energy flow to the photosynthetic reaction centers. Upon binding of FRP, OCP^R is converted to OCP^O and dissociates from PBs; however, the mode and site of OCP^R/FRP interactions remain elusive. Recently, we have introduced the purple OCP^W288A mutant as a competent model for the signaling state OCP^R (Sluchanko et al., Biochim Biophys Acta 1858:1–11, 2017). Here, we have utilized fluorescence labeling of OCP at its native cysteine residues to generate fluorescent OCP proteins for fluorescence correlation spectroscopy (FCS). Our results show that OCP^W288A has a 1.6(±0.4)-fold larger hydrodynamic radius than OCP^O, supporting the hypothesis of domain separation upon OCP photoactivation. Whereas the addition of FRP did not change the diffusion behavior of OCP^O, a substantial compaction of the OCP^W288A mutant and of the OCP apoprotein was observed. These results show that sufficiently stable complexes between FRP and OCP^W288A or the OCP apoprotein are formed to be detected by FCS. 1:1 complex formation with a micromolar apparent dissociation constant between OCP apoprotein and FRP was confirmed by size-exclusion chromatography. Beyond the established OCP/FRP interaction underlying NPQ cessation, the OCP apoprotein/FRP interaction suggests a more general role of FRP as a scaffold protein for OCP maturation.     

Yeast Prions Compared to Functional Prions and Amyloids

Saccharomyces cerevisiae is an occasional host to an array of prions, most based on self-propagating, self-templating amyloid filaments of a normally soluble protein. [URE3] is a prion of Ure2p, a regulator of nitrogen catabolism, while [PSI +] is a prion of Sup35p, a subunit of the translation termination factor Sup35p. In contrast to the functional prions, [Het-s] of Podospora anserina and [BETA] of yeast, the amyloid-based yeast prions are rare in wild strains, arise sporadically, have an array of prion variants for a single prion protein sequence, have a folded in-register parallel β-sheet amyloid architecture, are detrimental to their hosts, arouse a stress response in the host, and are subject to curing by various host anti-prion systems. These characteristics allow a logical basis for distinction between functional amyloids/prions and prion diseases. These infectious yeast amyloidoses are outstanding models for the many common human amyloid-based diseases that are increasingly found to have some infectious characteristics.