Author Correction: Jawsamycin exhibits in vivo antifungal properties by inhibiting Spt14/Gpi3-mediated biosynthesis of glycosylphosphatidylinositol.

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Jawsamycin exhibits in vivo antifungal properties by inhibiting Spt14/Gpi3-mediated biosynthesis of glycosylphosphatidylinositol.

Biosynthesis of glycosylphosphatidylinositol (GPI) is required for anchoring proteins to the plasma membrane, and is essential for the integrity of the fungal cell wall. Here, we use a reporter gene-based screen in Saccharomyces cerevisiae for the discovery of antifungal inhibitors of GPI-anchoring of proteins, and identify the oligocyclopropyl-containing natural product jawsamycin (FR-900848) as a potent hit. The compound targets the catalytic subunit Spt14 (also referred to as Gpi3) of the fungal UDP-glycosyltransferase, the first step in GPI biosynthesis, with good selectivity over the human functional homolog PIG-A.

FR171456 is a specific inhibitor of mammalian NSDHL and yeast Erg26p.

FR171456 is a natural product with cholesterol-lowering properties in animal models, but its molecular target is unknown, which hinders further drug development. Here we show that FR171456 specifically targets the sterol-4-alpha-carboxylate-3-dehydrogenase (Saccharomyces cerevisiae--Erg26p, Homo sapiens--NSDHL (NAD(P) dependent steroid dehydrogenase-like)), an essential enzyme in the ergosterol/cholesterol biosynthesis pathway. FR171456 significantly alters the levels of cholesterol pathway intermediates in human and yeast cells.

Nannocystin A: an Elongation Factor 1 Inhibitor from Myxobacteria with Differential Anti-Cancer Properties.

Cultivation of myxobacteria of the Nannocystis genus led to the isolation and structure elucidation of a class of novel cyclic lactone inhibitors of elongation factor 1. Whole genome sequence analysis and annotation enabled identification of the putative biosynthetic cluster and synthesis process. In biological assays the compounds displayed anti-fungal and cytotoxic activity.

4-Aminothiazolyl analogues of GE2270 A: antibacterial lead finding.

4-Aminothiazolyl analogues of the antibacterial natural product GE2270 A (1) were designed, synthesized, and evaluated for gram positive bacteria growth inhibition. The aminothiazole-based chemical template was evaluated for chemical stability, and its decomposition revealed a novel, structurally simplified, des-thiazole analogue of 1. Subsequent stabilization of the 4-aminothiazolyl functional motif was achieved and initial structure activity relationships defined.

Ribosomally synthesized thiopeptide antibiotics targeting elongation factor Tu.

We identified the thiomuracins, a novel family of thiopeptides produced by a rare-actinomycete bacterium typed as a Nonomuraea species, via a screen for inhibition of growth of the bacterial pathogen Staphylococcus aureus. Thiopeptides are a class of macrocyclic, highly modified peptides that are decorated by thiazoles and defined by a central six-membered heterocyclic ring system. Mining the genomes of thiopeptide-producing strains revealed the elusive biosynthetic route for this class of antibiotics.

Optimisation of protein expression and establishment of the Wave Bioreactor for Baculovirus/insect cell culture.

As the interest of research is beginning to shift from genomicsto proteomics the number of proteins to be expressed is rapidlyincreasing. To do so, well-established, high-level expressionsystems and rapid, cost-effective production means are needed. For addressing the latter, a novel cultivation system for recombinant cells, the Wave Bioreactortrade mark has recently becomeavailable.

Epothilones as lead structures for new anticancer drugs--pharmacology, fermentation, and structure-activity-relationships.

Epothilones (Epo's) A and B are naturally occurring microtubule-stabilizers, which inhibit the growth of human cancer cells in vitro at low nM or sub-nM concentrations. In contrast to taxol (paclitaxel, Taxol) epothilones are also active against different types of multidrug-resistant cancer cell lines in vitro and against multidrug-resistant tumors in vivo. Their attractive preclinical profile has made epothilones important lead structures in the search for improved cytotoxic anticancer drugs and Epo B (EPO906, patupilone) is currently undergoing Phase III clinical trials.

Gas-inducible product gene expression in bioreactors.

Inducible transgene expression technologies are of unmatched potential for biopharmaceutical manufacturing of unstable, growth-impairing and cytotoxic proteins as well as conditional metabolic engineering to improve desired cell phenotypes. Currently available transgene dosing modalities which rely on physical parameters or small-molecule drugs for transgene fine-tuning compromise downstream processing and/or are difficult to implement technologically. The recently designed gas-inducible acetaldehyde-inducible regulation (AIR) technology takes advantage of gaseous acetaldehyde to modulate product gene expression levels.

Argyrins, immunosuppressive cyclic peptides from myxobacteria. I. Production, isolation, physico-chemical and biological properties.

A group of cyclic peptides consisting of 8 amino acid residues, named argyrins A to H, were isolated from the culture broth of strains of the myxobacterium Archangium gephyra. Argyrin B was found to be a potent inhibitor of T cell independent antibody formation by murine B cells and strongly inhibited the two way murine mixed lymphocyte reaction. All argyrins had slight antibiotic activity, especially against Pseudomonas sp.

Sanglifehrins A, B, C and D, novel cyclophilin-binding compounds isolated from Streptomyces sp. A92-308110. I. Taxonomy, fermentation, isolation and biological activity.

A novel class of macrolides for which the name sanglifehrins is proposed, has been discovered from actinomycete strains based on their high affinity binding for cyclophilin A (CypA), an immunophilin originally identified as a cytosolic protein binding cyclosporin A (CsA). The sanglifehrins were produced by Streptomyces sp. A92-308110.

Expression, refolding, and autocatalytic proteolytic processing of the interleukin-1 beta-converting enzyme precursor.

The interleukin-1 beta-converting enzyme is a heterodimeric cysteine protease that is produced as a 45-kDa precursor. The full-length precursor form of the enzyme was expressed in Escherichia coli as insoluble inclusion bodies. Following solubilization and refolding of the 45-kDa protein, autoproteolytic conversion to a heterodimeric form containing 10- and 20-kDa subunits was observed.

A novel approach for high level production of a recombinant human parathyroid hormone fragment in Escherichia coli.

We describe a novel approach to the production in E. coli of a peptide fragment derived from the human parathyroid hormone (hPTH). The first 38 amino acids of hPTH were fused at the amino terminus to a derivative of the bacteriophage T4-encoded gp55 protein, and were expressed in the E.

Biosynthesis of the unusual amino acid (4R)-4-[(E)-2-butenyl]-4-methyl-L-threonine of cyclosporin A. Identification of 3(R)-hydroxy-4(R)-methyl-6(E)-octenoic acid as a key intermediate by enzymatic in vitro synthesis and by in vivo labeling techniques.

The biosynthesis of (4R)-4-[(E)-2-butenyl]-4-methyl-L-threonine (abbreviation: Bmt, systematic name: 2(S)-amino-3(R)-hydroxy-4(R)-methyl-6(E)-octenoic acid) is proposed to involve two principal phases: the formation of a polyketide backbone and a subsequent transformation process to the final product. Here we report on the identification of 3(R)-hydroxy-4(R)-methyl-6(E)-octenoic acid as the end product of the first phase. The primary indication of 3(R)-hydroxy-4(R)-methyl-6(E)-octenoic acid as the key intermediate in the proposed biosynthetic route came from in vivo labeling studies with [1-13C,18O2]acetate, demonstrating retention of 18O in the 3-hydroxy group.

Effect of specific growth rates on productivity in continuous open and partial cell retention animal cell bioreactors.

A clonal derivative of a transfectant of the SP2/0 myeloma cell line producing a chimeric monoclonal antibody was cultivated in both continuous open and continuous partially-closed bioreactors. Using an open system for the determination of kinetic parameters, we showed that the production of this chimeric mAb was growth associated. As such, the volumetric productivity increased linearly with increasing dilution rate up to the maximum dilution rate.

Kinetics of recombinant immunoglobulin production by mammalian cells in continuous culture.

A clonal derivative of a transfectant of the SP2/O myeloma cell line producing a chimeric monoclonal antibody was maintained in steady-state, continuous culture at dilution rates ranging from 0.21 to 1.04 day(-1).

Protein secondary structure determination by NMR. Application with recombinant human cyclophilin.

It is a unique trait of the NMR method for protein structure determination that a description of the polypeptide secondary structure can be obtained at an early stage and quite independently of the complete structure calculation. In this paper the procedures used for secondary structure determination are reviewed and placed in perspective relative to the other steps in a complete three-dimensional structure determination. As an illustration the identification of the regular secondary structure elements in human cyclophilin is described.

Fractional gastrointestinal transit time: intra- and interindividual variation.

The intra- and interindividual variation in the gastrointestinal transit times was measured in 12 healthy adult test subjects using a scintigraphic method with 111In marked single unit tablets. Test conditions were semi-standardized. The variation within the same individual was as large as the variation between different subjects regarding the transit times through both the small and the large intestines.