A novel β2-AR agonist,Higenamine, induces β-arrestin-biased signaling

Science China Life Sciences - The biased ligands in G protein-coupled receptors (GPCRs) have opened new avenues for developing safer and more effective drugs. However, the identification of such...     

Novel mutations involving βI-, βIIA-, or βIVB-tubulin isotypes with functional resemblance to βIII-tubulin in breast cancer

Tubulin is the target for very widely used anti-tumor drugs, including Vinca alkaloids, taxanes, and epothilones, which are an important component of chemotherapy in breast cancer and other malignancies. Paclitaxel and other tubulin-targeting drugs bind to the β subunit of tubulin, which is a heterodimer of α and β subunits. β-Tubulin exists in the form of multiple isotypes, which are differentially expressed in normal and neoplastic cells and differ in their ability to bind to drugs. Among them, the βIII isotype is overexpressed in many aggressive and metastatic cancers and may serve as a prognostic marker in certain types of cancer. The underpinning mechanisms accounting for the overexpression of this isotype in cancer cells are unclear. To better understand the role of β-tubulin isotypes in cancer, we analyzed over 1000 clones from 90 breast cancer patients, sequencing their β-tubulin isotypes, in search of novel mutations. We have elucidated two putative emerging molecular subgroups of invasive breast cancer, each of which involve mutations in the βI-, βIIA-, or βIVB isotypes of tubulin that increase their structural, and possibly functional, resemblance to the βIII isotype. A unifying feature of the first of the two subgroups is the mutation of the highly reactive C239 residue of βI- or βIVB-tubulin to L239, R239, Y239, or P239, culminating in probable conversion of these isotypes from ROS-sensitive to ROS-resistant species. In the second subgroup, βI, βIIA, and βIVB have up to seven mutations to the corresponding residues in βIII-tubulin. Given that βIII-tubulin has emerged as a pro-survival factor, overexpression of this isotype may confer survival advantages to certain cancer cell types. In this mini-review, we bring attention to a novel mechanism by which cancer cells may undergo adaptive mutational changes involving alternate β-tubulin isotypes to make them acquire some of the pro-survival properties of βIII-tubulin. These “hybrid” tubulins, combining the sequences and/or properties of two wild-type tubulins (βIII and either βI, βIIA, or βIVB), are novel isotypes expressed solely in cancer cells and may contribute to the molecular understanding and stratification of invasive breast cancer and provide novel molecular targets for rational drug development.     

Synthesis of disaccharide derivatives employing β-N-acetyl- d-hexosaminidase, β-d-galactosidase and β-d-glucuronidase

-N-Acetyl-d-hexosaminidase from Aspergillus oryzae catalysed the stereo- and regiospecific formation of the 6-O-benzylated disaccharide derivatives GalNAc1-3(6- OBn)Gal-SEt and GlcNAc1-3(6-OBn)Gal-SEt, which were obtained in transglycosylation reactions employing ethyl 6- O-benzyl-1-thio--d-galactopyranoside as acceptor. Preparative amounts of the chitobiose derivative GlcNAc1- 3GlcNAc-OPhNO2-p was prepared as well. - N-Acetyl-d-hexosaminidase from bovine testes catalysed the specific synthesis of GlcNAc1-3(6-OBn)GlcNH2-SEt and GalNAc1-3(6-OBn)GlcNH2-SEt, employing ethyl 2-amino-6-O-benzyl-2-deoxy-1-thio--d-glucopyranoside as acceptor. -d-Glucuronidase from E. coli was found to catalyse the formation of GlcA1-3(6-OBn)GlcNH2- SEt employing the same acceptor.     

Separation and properties of β-galactosidase, β-glucosidase, β-glucuronidase and n-acetyl-β-glucosaminidase from rat kidney

1. The activities of β-galactosidase, β-glucosidase, β-glucuronidase and N-acetyl-β-glucosaminidase from rat kidney have been compared when 4-methylumbelliferyl glycosides are used as substrates. 2. Separation by gel electrophoresis at pH7·0 indicated slow- and fast-moving components of rat-kidney β-galactosidase. 3. The fast-moving component is also associated with the total β-glucosidase activity and inhibition experiments indicate that a single enzyme species is responsible for both activities. 4. DEAE-cellulose chromatography and filtration on Sephadex gels suggests that the β-glucosidase component is a small acidic molecule, of molecular weight approx. 40000–50000, with optimum pH5·5–6·0 for β-galactosidase and β-glucosidase activities. 5. The major β-galactosidase component has low electrophoretic mobility, a calculated molecular weight of 80000 and optimum pH3·7.     

Eicosanoids, β-cell function, and diabetes

Arachidonic acid (AA) is metabolized by cyclooxygenase (COX), lipoxygenase (LOX), and cytochrome P450 (CYP) enzymes into eicosanoids, which are involved in diverse diseases, including type 1 and type 2 diabetes. During the last 30 years, evidence has been accumulated that suggests important functions for eicosanoids in the control of pancreatic β-cell function and destruction. AA metabolites of the COX pathway, especially prostaglandin E(2) (PGE(2)), appear to be significant factors to β-cell dysfunction and destruction, participating in the pathogenesis of diabetes and its complications. Several elegant studies have contributed to the sorting out of the importance of 12-LOX eicosanoids in cytokine-mediated inflammation in pancreatic β cells. The role of CYP eicosanoids in diabetes is yet to be explored. A recent publication has demonstrated that stabilizing the levels of epoxyeicosatrienoic acids (EETs), CYP eicosanoids, by inhibiting or deleting soluble epoxide hydrolase (sEH) improves β-cell function and reduces β-cell apoptosis in diabetes. In this review we summarize recent findings implicating these eicosanoid pathways in diabetes and its complications. We also discuss the development of animal models with targeted gene deletion and specific enzymatic inhibitors in each pathway to identify potential targets for the treatment of diabetes and its complications.     

Memory, synapse stability, and β-adducin

In this issue of Neuron, two studies by Pielage et?al. and Bednarek and Caroni suggest that the cytoskeleton regulator β-Adducin provides an activity-dependent switch controlling synapse disassembly and assembly at the Drosophila neuromuscular junction (NMJ) and the mouse hippocampus. In mice, the β-Adducin switch is required for the improvement of learning and memory induced by enriched environments.     

Chorein,the protein responsible for chorea-acanthocytosis,interacts with β-adducin and β-actin

Chorea-acanthocytosis (ChAc) is an autosomal, recessive hereditary disease characterized by striatal neurodegeneration and acanthocytosis, and caused by loss of function mutations in the vacuolar protein sorting 13 homolog A (VPS13A) gene. VPS13A encodes chorein whose physiological function at the molecular level is poorly understood. In this study, we show that chorein interacts with β-adducin and β-actin. We first compare protein expression in human erythrocyte membranes using proteomic analysis. Protein levels of β-adducin isoform 1 and β-actin are markedly decreased in erythrocyte membranes from a ChAc patient. Subsequent co-immunoprecipitation (co-IP) and reverse co-IP assays using extracts from chorein-overexpressing human embryonic kidney 293 (HEK293) cells, shows that β-adducin (isoforms 1 and 2) and β-actin interact with chorein. Immunocytochemical analysis using chorein-overexpressing HEK293 cells demonstrates co-localization of chorein with β-adducin and β-actin. In addition, immunoreactivity of β-adducin isoform 1 is significantly decreased in the striatum of gene-targeted ChAc-model mice. Adducin and actin are membrane cytoskeletal proteins, involved in synaptic function. Expression of β-adducin is restricted to the brain and hematopoietic tissues, corresponding to the main pathological lesions of ChAc, and thereby implicating β-adducin and β-actin in ChAc pathogenesis.     

Crystal structures of rare disaccharides, α-D-glucopyranosyl β-D-psicofuranoside, and α-D-galactopyranosyl β-D-psicofuranoside

The crystal structures of α-D-glucopyranosyl β-D-psicofuranoside and α-D-galactopyranosyl β-D-psicofuranoside were determined by a single-crystal X-ray diffraction analysis, refined to R(1)=0.0307 and 0.0438, respectively. Both disaccharides have a similar molecular structure, in which psicofuranose rings adopt an intermediate form between (4)E and (4)T(3). Unique molecular packing of the disaccharides was found in crystals, with the molecules forming a layered structure stacked along the y-axis.     

Synthesis of β-1, 3-Glucan and β-1, 3 and β-1, 4-Mixed Glucan from UDP-α-d-Glucose

A particulate enzyme preparation from Phaseolus aureus (mung bean) seedlings catalyzed the synthesis of a water insoluble β-1,3-glucan from UDP-α-d-glucose (UDPG) at high concentrations (0.4~20 mm) and an alkaline insoluble β-1,3 and β-1,4-mixed glucan from UDPG at a low concentration (8.5 µm).

Furthermore, the two kinds of β-glucan synthetases which were investigated with two reaction systems at high and low concentrations of UDPG had different properties in optimal pH, stability of enzyme activity, and metallic ion requirement.     

Soymorphins,Novel μ Opioid Peptides Derived from Soy β-Conglycinin β-Subunit,Have Anxiolytic Activities

Based on the amino acid sequence YPFV found in the soy β-conglycinin β-subunit, which is common to an opioid peptide human β-casomorphin-4, peptides YPFVV, YPFVVN, and YPFVVNA were synthesized according to their primary structure. On guinea pig ileum (GPI) assay, they showed opioid activity (IC₅₀ = 6.0, 9.2 and 13 μM respectively) more potent than human β-casomorphins, and were named soymorphins-5, -6, and -7, respectively. Their opioid activities on mouse vas deferens (MVD) assay were less potent than on GPI assay, suggesting that they are selective for the μ opioid receptor. Human β-casomorphin-4 and soymorphin-5 were released from the soy 7S fraction (β-conglycinin) by the action of gastrointestinal proteases. Soymorphins-5, -6, and -7 had anxiolytic activities after oral administration at doses of 10–30 mg/kg in the elevated plus-maze test in mice.     

Ultrastructural localization of corticotropin, β-lipotropin,and αand β-endorphin in the porcine anterior pituitary

Summary The peroxidase-antiperoxidase immunocytochemical technique was used to identify the ACTH/endorphin cells in the porcine pituitary at the ultrastructural level and to determine the precise subcellular localization of the pro-ACTH/endorphin fragments. The cells display different aspects: 1) large, regular shapes with numerous and large secretory granules; 2) small, irregular and angular shapes with small granules aligned along the periphery of the cell; and 3) intermediate forms. The presence of and -endorphin not only in the same cells but also in the same secretory granules that contain ACTH and -LPH clearly indicates that both the precursor or its fragments and the abovementioned peptides are stored in the same granules and released simultaneously by the corticotropic cells. The presence of FSH in some corticotropic cells is also discussed.Abbreviations used in this Article ACTH corticotropin - -MSH -melanotropin (ACTH I–I3) - CLIP corticotropin-like intermediate lobe peptide (ACTH 18–39) - -LPH -lipotropin - -MSH -melanotropin (-LPH 41–58); -endorphin (-LPH 61–91); -endorphin (-LPH 61–76)     

Characterization of β-Lactotensin,a Bioactive Peptide Derived from Bovine β-Lactoglobulin,as a Neurotensin Agonist

β-Lactotensin (β-LT: His-Ile-Arg-Leu) is an ileum-contracting peptide derived from residues No. 146-149 of bovine β-lactoglobulin. The ileum-contracting activity of β-LT was blocked by the NT₁ antagonist SR48692. β-LT was selective for the neurotensin NT₂ receptor while neurotensin was selective for the NT₁ receptor. β-LT is the first natural ligand showing selectivity for the NT₂ receptor. β-LT showed hypertensive activity after intravenous administration at a dose of 30 mg/kg in conscious rats, while neurotensin showed hypotensive activity. The hypertensive activity of β-LT was blocked by levocabastine (1 mg/kg, i.v.), an NT₂ antagonist. SR48692, which blocked the hypotensive activity of neurotensin, had no effect on the hypertensive activity of β-LT. These results suggest that the hypertensive activity of β-LT is mediated by the NT₂ receptor. It was concluded that the NT₁ and NT₂ receptors mediate the opposite effect on blood pressure.     

The β-lactam antibiotics: past, present, and future

The discovery and development of the -lactam antibiotics are among the most powerful and successful achievements of modern science and technology. Since Fleming's accidental discovery of the penicillin-producing mold, seventy years of steady progress has followed, and today the -lactam group of compounds are the most successful example of natural product application and chemotherapy. Following on the heels of penicillin production by Penicillium chrysogenum came the discoveries of cephalosporin formation by Cephalosporium acremonium, cephamycin, clavam and carbapenem production by actinomycetes, and monocyclic -lactam production by actinomycetes and unicellular bacteria. Each one of these groups has yielded medically-useful products and has contributed to the reduction of pain and suffering of people throughout the world. Research on the microbiology, biochemistry, genetics and chemistry of these compounds have continued up to the present with major contributions being made by both individual and collaborative groups from industry and academia. The discovery of penicillin not only led to the era of the wonder drugs but provided the most important antibiotics available to medicine. Continued efforts have resulted in the improvement of these compounds with respect to potency, breadth of spectrum, activity against resistant pathogens, stability and pharmacokinetic properties. On the research front, major advances are being made on structural and regulatory biosynthetic genes and metabolic engineering of the pathways involved. New semisynthetic compounds especially those designed to combat resistance development are being examined in the clinic, and unusual non-antibiotic activities of these compounds are being pursued. Although seventy years of age, the -lactams are not yet ready for retirement.     

Barley β-Galactosidase: Structure, Function, Heterogeneity, and Gene Origin

Barley (Hordeum vulgare) β-galactosidase is composed of a large (45 kDa) and a small (33 kDa) polypeptide. N-terminal sequencing of the polypeptides and antibody reactivity data place the barley enzyme and heterodimeric plant β-galactosidases from jack bean, maize, and wheat in family 35 of the glycosyl hydrolases. Sequence analysis indicates the existence of a subfamily of genes coding for polypeptide precursors that are cleaved to produce the two subunits in heterodimeric β-galactosidases. The heterogeneity of the barley holoenzyme is related, but not restricted, to the N-glycosylation of the small polypeptide. Both polypeptides are essential for the catalytic activity of the enzyme.     

Potent transglutaminase inhibitors,aryl β-aminoethyl ketones

Aryl β-aminoethyl ketones were discovered as potent inhibitors of tissue transglutaminase. Heteroaryl-like thiophene groups and N-benzyl N-t-butyl aminoethyl group are critical to the strong inhibitory activity of aryl β-aminoethyl ketones.     

Potent transglutaminase inhibitors, dithio β-aminoethyl ketones

Potent transglutaminase inhibitors were obtained from disulfide compounds, cystamine, dimethyl cystine, and dimethyl homocystine. The disulfide bond and thiophene ring play an important role in inhibitory activity of synthesized aryl β-amino ketones.     

Production, Characterization, and Properties of β-Glucosidase and β-Xylosidase from a Strain of Aureobasidium sp.

-Glucosidase and -xylosidase production by a yeastlike Aureobasidium sp. was carried out during solid-state and submerged fermentation using different carbon sources and crude enzymes were characterized. -Glucosidase and -xylosidase exhibited optimum activities at pH 2.0–2.5 and 3.0, respectively. These enzymes had the maximum activities at 65°C and were stable in a wide pH range and at high temperatures.     

Conformational Analysis of the β-amyloid Peptide Fragment, β(12–28)

Abstract

NMR and CD spectroscopy have been used to examine the conformation of the peptide, β(12–28), (VHHQKLVFFAEDVGSNK) in aqueous and 60% TFE/40% H₂0 solution at pH 2.4. In 60% TFE solution, the peptide is helical as confirmed by the CD spectrum and by the pattern of the NOE cross peaks detected in the NOESY spectrum of the peptide. In aqueous solution, the peptide adopts a more extended and flexible conformation. Broadening of resonances at low temperature, temperature-dependent changes in the chemical shifts of several of the CH_α resonances and the observation of a number of NOE contacts between the hydrophobic side-chain protons of the peptide are indicative of aggregation in aqueous solution. The behavior of β(12–28) in 60% TFE and in aqueous solution are consistent with the overall conformation and aggregation behavior reported for the larger peptide fragment, β(1–28) and the parent β-amyloid peptide.     

Conformational Analysis of the β-amyloid Peptide Fragment, β(12-28)

Abstract NMR and CD spectroscopy have been used to examine the conformation of the peptide, β(12-28), (VHHQKLVFFAEDVGSNK) in aqueous and 60% TFE/40% H(2)0 solution at pH 2.4. In 60% TFE solution, the peptide is helical as confirmed by the CD spectrum and by the pattern of the NOE cross peaks detected in the NOESY spectrum of the peptide. In aqueous solution, the peptide adopts a more extended and flexible conformation. Broadening of resonances at low temperature, temperature-dependent changes in the chemical shifts of several of the CH(α) resonances and the observation of a number of NOE contacts between the hydrophobic side-chain protons of the peptide are indicative of aggregation in aqueous solution. The behavior of β(12-28) in 60% TFE and in aqueous solution are consistent with the overall conformation and aggregation behavior reported for the larger peptide fragment, β(1-28) and the parent β-amyloid peptide.