Influence of cholesterol on tetraphenylborate transport through rat brain lipid artificial membranes

Summary The influence of cholesterol on TΦB⁻ transport through rat brain lipid model membranes has been studied by performing conductance studies. As a model system was used a rat brain lipid filter supported membrane. The weight percent of cholesterol as referred to total lipids was varied in a wide range. Cholesterol produced an increase of the conductance of membranes induced by the lipophilic anion tetraphenylborate. Moroever, an increase in the cholesterol mole fraction had a strong influence on steady-state I–V characteristics of rat brain lipid model membranes. To speed up publication, the authors of this paper have agreed to not receive the proofs for correction.     

Two‐fold Bioorthogonal Derivatization by Different Formylglycine‐Generating Enzymes

Formylglycine‐generating enzymes are of increasing interest in the field of bioconjugation chemistry. They catalyze the site‐specific oxidation of a cysteine residue to the aldehyde‐containing amino acid C^α‐formylglycine (FGly). This non‐canonical residue can be generated within any desired target protein and can subsequently be used for bioorthogonal conjugation reactions. The prototypic formylglycine‐generating enzyme (FGE) and the iron‐sulfur protein AtsB display slight variations in their recognition sequences. We designed specific tags in peptides and proteins that were selectively converted by the different enzymes. Combination of the different tag motifs within a single peptide or recombinant protein enabled the independent and consecutive introduction of two formylglycine residues and the generation of heterobifunctionalized protein conjugates.     

On‐Line Polyketide Cyclization into Diverse Medium‐Sized Lactones by a Specialized Ketosynthase Domain

Ketosynthase (KS) domains of modular type I polyketide synthases (PKSs) typically catalyze the Claisen condensation of acyl and malonyl units to form linear chains. In stark contrast, the KS of the rhizoxin PKS branching module mediates a Michael addition, which sets the basis for a pharmacophoric δ‐lactone moiety. The precise role of the KS was evaluated by site‐directed mutagenesis, chemical probes, and biotransformations. Biochemical and kinetic analyses helped to dissect branching and lactonization reactions and unequivocally assign the entire sequence to the KS. Probing the range of accepted substrates with diverse synthetic surrogates in vitro, we found that the KS tolerates defined acyl chain lengths to produce five‐ to seven‐membered lactones. These results show that the KS is multifunctional, as it catalyzes β‐branching and lactonization. Information on the increased product portfolio of the unusual, TE‐independent on‐line cyclization is relevant for synthetic biology approaches.     

Comparative studies of immobilized chiral stationary phases based on polysaccharide derivatives for enantiomeric separation of 15 azole compounds

Immobilized polysaccharide‐based columns showed excellent enantioselectivity in normal phase separation mode. In this work, enantioseparation abilities of four immobilized polysaccharide‐derived chiral stationary phases (Chiralpak IA, Chiralpak IB, Chiralpak IC, and Chiralpak ID) toward 15 azole compounds were evaluated. Separation was carried out using n‐hexane as mobile phase with ethanol, 1‐propanol, 1‐butanol, and 2‐propanol as modifiers. And twelve compounds have achieved baseline separation with the resolutions ranging between 2.05 and 21.73. The enantioseparation on the four polysaccharide‐based chiral columns using different alcohol modifiers was compared. In general, the best separation performance was identified as Chiralpak IC, which was able to resolve 11 compounds to baseline and two partially under the screening conditions. Separation on Chiralpak IB was not satisfactory, because only four compounds were baseline separated.     

“Head‐to‐Middle” and “Head‐to‐Tail” cis‐Prenyl Transferases: Structure of Isosesquilavandulyl Diphosphate Synthase

We report the first X‐ray crystallographic structure of the “head‐to‐middle” prenyltransferase, isosesquilavandulyl diphosphate synthase, involved in biosynthesis of the merochlorin class of antibiotics. The protein adopts the ζ or cis‐prenyl transferase fold but remarkably, unlike tuberculosinol adenosine synthase and other cis‐prenyl transferases (e.g. cis‐farnesyl, decaprenyl, undecaprenyl diphosphate synthases), the large, hydrophobic side chain does not occupy a central hydrophobic tunnel. Instead, it occupies a surface pocket oriented at 90° to the hydrophobic tunnel. Product chain‐length control is achieved by squeezing out the ligand from the conventional allylic S1 binding site, with proton abstraction being achieved using a diphosphate‐Asn‐Ser relay. The structures revise and unify our thinking as to the mechanism of action of many other prenyl transferases and may also be of use in engineering new merochlorin‐class antibiotics.     

Solvent‐Free Enzyme Activity: Quick,High‐Yielding Mechanoenzymatic Hydrolysis of Cellulose into Glucose

Mechanochemistry enables enzymatic cleavage of cellulose into glucose without bulk solvents, acids, other aggressive reagents, or substrate pre‐treatment. This clean mechanoenzymatic process (coined RAging) is also directly applicable to biomass, avoids many limitations associated with the use of cellulases, and produces glucose concentrations greater than three times that obtained by conventional methods.     

Imaging of Enzyme Activity by Electron Paramagnetic Resonance: Concept and Experiment Using a Paramagnetic Substrate of Alkaline Phosphatase

Enzyme activities are well established biomarkers of many pathologies. Imaging enzyme activity directly in vivo may help to gain insight into the pathogenesis of various diseases but remains extremely challenging. In this communication, we report the use of EPR imaging (EPRI) in combination with a specially designed paramagnetic enzymatic substrate to map alkaline phosphatase activity with a high selectivity, thereby demonstrating the potential of EPRI to map enzyme activity.     

Tailormade Polysaccharides with Defined Branching Patterns: Enzymatic Polymerization of Arabinoxylan Oligosaccharides

The heterogeneous nature of non‐cellulosic polysaccharides, such as arabinoxylan, makes it difficult to correlate molecular structure with macroscopic properties. To study the impact of specific structural features of the polysaccharides on crystallinity or affinity to other cell wall components, collections of polysaccharides with defined repeating units are required. Herein, a chemoenzymatic approach to artificial arabinoxylan polysaccharides with systematically altered branching patterns is described. The polysaccharides were obtained by glycosynthase‐catalyzed polymerization of glycosyl fluorides derived from arabinoxylan oligosaccharides. X‐ray diffraction and adsorption experiments on cellulosic surfaces revealed that the physicochemical properties of the synthetic polysaccharides strongly depend on the specific nature of their substitution patterns. The artificial polysaccharides allow structure–property relationship studies that are not accessible by other means.     

Inclusion chemistry for the modeling of heme proteins

Early attention to the modeling of heme proteins is enhancing the understanding of biochemistry. Those studies are also contributing to the development of techniques for the modeling of still more intricate, multifunctional, variously selective natural systems. Selectivity in simple systems may involve the molecular capability to bind only one of a family of related species or it may mean the ability to select and control one of a number of possible functions of a given bound species. Complicated systems simultaneously combine the two kinds of simple selectivities for two or more different classes of guest, often with synergistic interrelationships. The subject is developed around examples of binary, tertiary, and quarternary complexes designed to model the behavior of monooxygenases.