Dietmar G. Schmid

Molecular Characterization of Lantibiotic-synthesizing Enzyme EpiD Reveals a Function for Bacterial Dfp Proteins in Coenzyme A Biosynthesis

The lantibiotic-synthesizing flavoprotein EpiD catalyzes the oxidative decarboxylation of peptidylcysteines to peptidyl-aminoenethiols. The sequence motif responsible for flavin coenzyme binding and enzyme activity is conserved in different proteins from all kingdoms of life. Dfp proteins of eubacteria and archaebacteria and salt tolerance proteins of yeasts and plants belong to this new family of flavoproteins. The enzymatic function of all these proteins was not known, but our experiments suggested that they catalyze a similar reaction like EpiD and/or may have similar substrates and are homododecameric flavoproteins. We demonstrate that the N-terminal domain of the Escherichia coli Dfp protein catalyzes the decarboxylation of (R)-4′-phospho-N-pantothenoylcysteine to 4′-phosphopantetheine. This reaction is essential for coenzyme A biosynthesis.

Fluorescent and Electroactive Cyclic Assemblies from Perylene Tetracarboxylic Acid Bisimide Ligands and Metal Phosphane Triflates

Tetraaryloxy-substituted perylene tetracarboxylic acid bisimides with one or two 4-pyridyl receptor substituents at the imide functionality were synthesized and employed in transition metal directed self-assembly with Pd(II) and Pt(II) phosphane triflates. Upon mixing of the components, quantitative formation of functional molecular square-type complexes containing four dye molecules and model complexes of a 2:1 (perylene bisimide ligand:transition metal ion) stoichiometry was observed. The isolated metallosupramolecular squares were characterized by 1H and 31P [1H] NMR spectroscopy as well as conventional electrospray ionization (ESI) and ESI-FTICR mass spectrometry, which gave evidence for the structure and the high stability of these giant cyclic dye assemblies (molecular weight (3a) 8172, Pt-Pt corner diagonal ca. 3.4 nm). Studies of the optical absorption and fluorescence properties and the electrochemistry and spectroelectrochemistry of both the perylene bisimide ligands and the perylene bisimide metal complexes show that Pt(II) coordination does not interfere with the optical and electrochemical properties of the perylene bisimide ligands; this gives squares with high fluorescence quantum yields (phiF (3a)=0.88) and three fully reversible redox couples. The latter could be unambiguously related to quantitative formation of perylene bisimide radical cations (E1/2 = +0.93 V vs. Fc/Fc+), radical anions (E1/2= - 1.01 V vs. Fc/Fc+), and dianions (E1/2 = -1.14 V vs. Fc/Fc+); these redox reactions change the charge state of the cyclic assembly from +12 to zero. In contrast, Pd(II) coordination influenced the electrochemical properties of the assembly because of an irreversible palladium reduction at E1/2= -1.15 V versus Fc/Fc+. Finally, dynamic ligand exchange processes between different metallosupramolecular assemblies were investigated by multinuclear NMR and electrospray mass spectrometry. These studies confirmed the reversible nature of the pyridine-Pt(II)/Pd(II) coordination process.

The Flavoprotein MrsD Catalyzes the Oxidative Decarboxylation Reaction Involved in Formation of the Peptidoglycan Biosynthesis Inhibitor Mersacidin

The lantibiotic mersacidin inhibits peptidoglycan biosynthesis by binding to the peptidoglycan precursor lipid II. Mersacidin contains an unsaturated thioether bridge, which is proposed to be synthesized by posttranslational modifications of threonine residue +15 and the COOH-terminal cysteine residue of the mersacidin precursor peptide MrsA. We show that the flavoprotein MrsD catalyzes the oxidative decarboxylation of the COOH-terminal cysteine residue of MrsA to an aminoenethiol residue. MrsD belongs to the recently described family of homo-oligomeric flavin-containing Cys decarboxylases (i.e., the HFCD protein family). Members of this protein family include the bacterial Dfp proteins (which are involved in coenzyme A biosynthesis), eukaryotic salt tolerance proteins, and further oxidative decarboxylases such as EpiD. In contrast to EpiD and Dfp, MrsD is a FAD and not an FMN-dependent flavoprotein. HFCD enzymes are characterized by a conserved His residue which is part of the active site. Exchange of this His residue for Asn led to inactivation of MrsD. The lantibiotic-synthesizing enzymes EpiD and MrsD have different substrate specificities.

FTICR-Mass Spectrometry for High-Resolution Analysis in Combinatorial Chemistry

The diversity of compound collections required for finding lead structures in pharmaceutical research can be provided by means of combinatorial organic chemistry. The resultant enormous number of single compounds but also of compound mixtures represents a challenge for the analyst. With the introduction of Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS or FT-MS), a new and, as yet, not widespread mass spectrometric technique (a means of analysis of such compound libraries with a very high mass resolution) high mass accuracy and high sensitivity has become available. Moreover, in combination with electrospray ionization (ESI), not only high-throughput measurements via flow-injection analysis (FIA) but also coupling with separation techniques such as high-performance liquid chromatography (HPLC) or capillary electrophoresis (CE) is possible. Structural verification by way of decomposing ions (MS(n); n > or = 2) using a variety of different dissociation techniques can be performed by FTICR-MS. This is the first review specifically covering applications of FTICR-MS in the field of combinatorial chemistry.

Alkylating Polymers: Resin-Released Carbenium Ions as Versatile Reactive Intermediates in Polymer-Assisted Solution-Phase Synthesis

Solid-supported diazoalkane analogues for the smooth and clean esterifications of carboxylic acids [Eq. (1)] as well as of complex compound mixtures: the alkylating polymers presented here were synthesized as solid-supported 3-alkyl-1-aryltriazenes and are capable of releasing carbenium ions following acidic activation.

Basidiochrome – A Novel Siderophore of the Orchidaceous Mycorrhizal Fungi Ceratobasidium and Rhizoctonia spp.

A novel trishydroxamate siderophore, named basidiochrome, was isolated as the principal siderophore from low-iron culture filtrates of Ceratobasidium and Rhizoctonia species which are known as mycorrhizal fungi associated with orchid roots. Ion-exchange chromatography and preparative HPLC yielded a pure compound which contained two components according to GC–MS analysis: l-N5-hydroxy-ornithine and 3-methyl-2-cis-pentenedioic acid (3-methyl-cis-glutaconic acid). FTICR-ESI-MS of both the iron-free and ferric form indicated an elemental composition of C33H47N6O16Fe (MW = 839) for the ferric form of basidiochrome. The connectivity was further elucidated by 2D-NMR techniques (HSQC, HMBC, COSY, NOESY) indicating that basidiochrome is a novel linear tripeptide consisting of three l-N5-hydroxy-ornithines each linked to 3-methyl-2-cis-pentenedioic acid residues.

Biaryl-Bridged Lipopeptides from Streptomyces sp. TÜ 6075

HPLC-DAD screening for new metabolites [1] from Streptomyces sp. TU 6075 revealed two series of compounds composed of five colorless and seven yellow fractions. For structure elucidation, ES-FTICR-MS, chiral GC-MS, Edman sequencing, and NMR spectroscopy were used. The isolated compounds, termed arylomycins, represent the first example of lipopeptides containing biaryl-bridged amino acids. The majority of the arylomycins exhibit antimicrobial activity against Gram-positive bacteria.

Solid-Supported Triazenes as Alkylating Polymers Employed for the Versatile Derivatization of Peptides

Recently, we have shown polymer-bound l-aryl-3-alkyl-triazenes to be versatile tools for the smooth esterification of various carboxylic acids [1], including peptides and products of combinatorial synthesis [2]. The triazenes are prepared in a 4-step synthesis on Merrifield resin. They react with carboxylic acids under loss of nitrogen and release of highly reactive carbenium ions forming the corresponding esters (Scheme 1).

Deamidation within a γ-Gliadin-Derived Peptide Enhances Its Recognition by Serum Antibodies of CD Patients

Coeliac disease (CD) or gluten sensitive enteropathy is an autoimmune disease mainly affecting the proximal small intestine [1]. It is caused in susceptible persons by the ingestion of certain cereal proteins, i.e. gliadins and glutenins from wheat. The intestinal mucosa shows characteristic lesions resulting in malabsorption of nutrients.