Synthesis and structural study of a fully protected α-C-galactosyl model dipeptide

Summary The synthesis of three N-alkyl-6,7-dideoxy-1,2:3,4-di-O-isopropylidene-7-[(alkyl-carbonyl)amino]-L-glycero-α-D-galacto-octopyranuronamides6a-c, analogous model dipeptides containing two amide groups connected to the α-carbon bearing the fully protected galactose as a side chain, has been realized with the aim of determining the conformational influence of the galactosyl moiety on the peptide backbone. Molecular modeing of6a, X-ray crystallography of6c and IR and NMR experiments on6a-c in organic solvents show that the carbohydrate ring assumes a twist boat conformation. In non-polar organic solvents, the NH of the left amide group interacts with one ketal oxygen of the galactosyl group.     

Temporal changes of mechanical signals and extracellular composition in human intervertebral disc during degenerative progression

In this study, a three-dimensional finite element model was used to investigate the changes in tissue composition and mechanical signals within human lumbar intervertebral disc during the degenerative progression. This model was developed based on the cell-activity coupled mechano-electrochemical mixture theory. The disc degeneration was simulated by lowering nutrition levels at disc boundaries, and the temporal and spatial distributions of the fixed charge density, water content, fluid pressure, Von Mises stress, and disc deformation were analyzed. Results showed that fixed charge density, fluid pressure, and water content decreased significantly in the nucleus pulposus (NP) and the inner to middle annulus fibrosus (AF) regions of the degenerative disc. It was found that, with degenerative progression, the Von Mises stress (relative to that at healthy state) increased within the disc, with a larger increase in the outer AF region. Both the disc volume and height decreased with the degenerative progression. The predicted results of fluid pressure change in the NP were consistent with experimental findings in the literature. The knowledge of the variations of temporal and spatial distributions of composition and mechanical signals within the human IVDs provide a better understanding of the progression of disc degeneration.     

A unique hairpin-type tail-anchored SNARE starts to solve a long-time puzzle

Macroautophagy mediates recycling of intracellular material by a multistep pathway, ultimately leading to the fusion of closed double-membrane structures, called autophagosomes, with the lysosome. This event ensures the degradation of the autophagosome content by lysosomal proteases followed by the release of macromolecules by permeases and, thus, it accomplishes the purpose of macroautophagy (hereafter referred to as autophagy). Because fusion of unclosed autophagosomes (i.e., phagophores) with the lysosome would fail to degrade the autophagic cargo, this critical step has to be tightly controlled. Yet, until recently, little was known about the regulation of this event and the factors orchestrating it. A punctum in this issue highlights the recent paper by Noboru Mizushima and his collaborators that answered the question of how premature fusion of phagophores with the lysosome is prevented prior to completion of autophagosome closure.     

6,7-Dichloroquinoxaline-2,3-Dione is a Competitive Antagonist Specific to Strychnine-Insensitive [3H]Glycine Binding Sites on the N-Methyl-D-Aspartate Receptor Complex

Multiple binding sites on the N-methyl-D-aspartate (NMDA) receptor complex were examined using rat brain synaptic membranes treated with Triton X-100. Binding of [3H](+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imi ne ([3H]MK-801), a noncompetitive NMDA antagonist, in the presence of 10 microM L-glutamate not only was inhibited by different types of antagonists, such as 6,7-dichloro-3-hydroxy-2-quinoxaline-carboxylate, 7-chlorokynurenate, and 6,7-dichloroquinoxaline-2,3-dione (DCQX), but also was abolished by non-NMDA antagonists, including 6-cyano-7-nitroquinoxaline-2,3-dione and 6,7-dinitroquinoxaline-2,3-dione. The inhibition of [3H]MK-801 binding by these compounds was invariably reversed or attenuated by addition of 10 microM glycine. Among these novel antagonists with an inhibitory potency on [3H]MK-801 binding, only DCQX abolished [3H]glycine binding without inhibiting [3H]glutamate and [3H](+-)-3-(2-carboxypiperazine-4-yl)propyl-1-phosphonate bindings. Other antagonists examined were all effective as displacers of the latter two bindings. These results suggest that DCQX is an antagonist highly selective to the strychnine-insensitive glycine binding sites with a relatively high affinity.     

A current view on Tau protein phosphorylation in Alzheimer's disease

The functions of the neuronal microtubule-associated protein Tau in the central nervous system are regulated by manifold posttranslational modifications at more than 50 sites. Tau in healthy neurons carries multiple phosphate groups, mostly in its microtubule assembly domain. Elevated phosphorylation and aggregation of Tau are widely considered pathological hallmarks in Alzheimer’s disease (AD) and other tauopathies, triggering the quest for Tau posttranslational modifications in the disease context. However, the phosphorylation patterns of physiological and pathological Tau are surprisingly similar and heterogenous, making it difficult to identify specific modifications as therapeutic targets and biomarkers for AD. We present a concise summary of - and view on - important previous and recent advances in Tau phosphorylation analysis in the context of AD.     

An Exclusive Increase in the Concentration of ATP as a Result of Osmotic Stress in Escherichia coli B

The ATP level was exclusively increased among the nucleotides in Escherichia coli under osmotic stress, concomitant with a decrease of guanosine phosphates levels. The profile of ATP formation was different from that resulting from chemicals. E. coli contains a specific system to enhance the ATP level and it might be possible that the source of the increased ATP is guanosine phosphates.     

Mixture design of starchy substrates hydrolysis by an immobilized glucoamylase from Aspergillus brasiliensis

Starch has great importance in human diet, since it is a heteropolymer of plants, mainly found in roots, as potato, cassava and arrowroots. This carbohydrate is composed by a highly-branched chain: amylopectin; and a linear chain: amylose. The proportion between the chains varies according to the botanical source. Starch hydrolysis is catalyzed by enzymes of the amilolytic system, named amylases. Among the various enzymes of this system, the glucoamylases (EC 3.2.1.3 glucan 1,4-alpha-glucosidases) are the majority because they hydrolyze the glycosidic linkages at the end of starch chains releasing glucose monomers. In this work, a glucoamylase secreted in the culture medium, by the ascomycete Aspergillus brasiliensis, was immobilized in Dietilaminoetil Sepharose-Polyethylene Glycol (DEAE-PEG), since immobilized biocatalysts are more stable in long periods of hydrolysis, and can be recovered from the final product and reused for several cycles. Glucoamylase immobilization has shown great thermal stability improvement over the soluble enzyme, reaching 66% more activity after 6?h at 60?°C, and 68% of the activity after 10 hydrolysis cycles. A simplex centroid experimental mixture design was applied as a tool to characterize the affinity of the immobilized enzyme for different starchy substrates. In assays containing several proportions of amylose, amylopectin and starch, the glucoamylase from A. brasiliensis mainly hydrolyzed the amylopectin chains, showing to have preference by branched substrates.     

Effective cancer therapy based on selective drug delivery into cells across their membrane using receptor-mediated endocytosis

Cancer is one of the major causes of death globally. The current treatment options are insufficient, leading to unmet medical needs in cancer treatment. Off-target side effects, multidrug resistance, selective distribution to cancerous tissues, and cell membrane permeation of anti-cancer agents are critical problems to overcome. There is a method to solve these problems by using receptor-mediated endocytosis (RME). It is well known that proteins such as integrin, HER2, EGFR, or other cancer biomarkers are specifically overexpressed on the surface of target cancer cells. By taking advantage of such specific receptors, payloads can be transported into cells through endocytosis using a conjugate composed of the corresponding ligands connected to the payloads by an appropriate linker. After RME, the payloads released by endosomal escape into the cytoplasm can exhibit the cytotoxic activity against cancer cells. Cell-penetrating peptides (CPPs), tumor-homing peptides (THPs), and monoclonal antibodies (mAbs) are utilized as ligands in this system. Antibody drug conjugates (ADCs) based on RME have already been used to cure cancer. In addition to the canonical conjugate method, nanocarriers for spontaneous accumulation in cancer tissue due to enhanced permeability and retention (EPR) effect are extensively used. In this review, I introduce the possibilities and advantages of drug design and development based on RME for the treatment of cancer.     

Preferential exclusion of hybrids in mixed pollinations between oilseed rape (Brassica napus) and weedy B. campestris (Brassicaceae)

In most experimental hybridizations between oilseed rape (Brassica napus) and weedy B. campestris, either intra- or interspecific pollen has been applied to individual flowers. Under field conditions, however, stigmas will often receive a mixture of the two types of pollen, thereby allowing for competition between male gametophytes and/or seeds within pods. To test whether competition influences the success of hybridization, pollen from the two species was mixed in different proportions and applied to stigmas of both species. The resulting seeds were scored for paternity by isozyme and randomly amplified polymorphic DNA analysis. Using data on the proportion of fully developed seeds and the proportion of these seeds that were hybrids, a statistical model was constructed to estimate the fitness of conspecific and heterospecific pollen and the survival of conspecific and heterospecific zygotes to seeds. B. campestris pollen in B. napus styles had a significantly lower fitness than the conspecific pollen, whereas no difference between pollen types was found in B. campestris styles. Hybrid zygotes survived to significantly lower proportions than conspecific zygotes in both species, with the lowest survival of hybrid zygotes in B. napus pods. This is in contrast to the higher survival of hybrid seeds in B. napus than in B. campestris pods when pollinations are made with pure pollen. Altogether, the likelihood of a foreign pollen grain producing a seed was much lower on B. napus than on B. campestris. In addition, pods on B. napus developed to a lower extent the more heterospecific pollen was in the mix, whereas this had no effect on B. campestris.