Palladium-Catalyzed Synthesis of -Trifluoromethyl Benzylic Amines via Fluoroarylation of -Difluoro-2-azadienes Enabled by Phosphine-Catalyzed Formation of an Azaallyl-Silver Intermediate.

We report the synthesis of -trifluoromethyl benzylic amines through the vicinal fluoroarylation of difluoro-2-azadienes. Our studies indicate that XPhos plays an important role as a phase transfer catalyst that promotes the addition of AgF to the difluoroazadiene, generating an trifluoromethyl azaallyl-silver intermediate that we have characterized by NMR spectroscopy. This intermediate likely transmetallates to Pd, coupling several aryl iodides to deliver products in up to 90% yield.

Formal Synthesis of (+)-Laurencin by Gold(I)-Catalyzed Intramolecular Dehydrative Alkoxylation.

8-Membered cyclic ethers are found in a wide range of natural products; however, they are challenging synthetic targets due to enthalpic and entropic barriers. The gold(I)-catalyzed intramolecular dehydrative alkoxylation of ω-hydroxy allylic alcohols was explored to stereoselectively construct α,α'-cis-oxocenes and further applied in a formal synthesis of (+)-laurencin. The gold(I)-catalyzed intramolecular dehydrative alkoxylation may constitute an alternative method for the synthesis of molecular building blocks and natural products that contain highly functionalized 8-membered cyclic ethers.

Identification of a cyclic nucleotide as a cryptic intermediate in molybdenum cofactor biosynthesis.

The molybdenum cofactor (Moco) is a redox cofactor found in all kingdoms of life, and its biosynthesis is essential for survival of many organisms, including humans. The first step of Moco biosynthesis is a unique transformation of guanosine 5'-triphosphate (GTP) into cyclic pyranopterin monophosphate (cPMP). In bacteria, MoaA and MoaC catalyze this transformation, although the specific functions of these enzymes were not fully understood.

A phosphoethanolamine-modified glycosyl diradylglycerol in the polar lipids of Clostridium tetani.

The polar lipids of the anaerobic bacterium Clostridium tetani, the causative agent of tetanus, have been examined by two-dimensional thin layer chromatography, ESI mass spectrometry, and NMR spectroscopy. Plasmalogen and di- and tetra-acylated species of phosphatidylethanolamine, phosphatidylglycerol, cardiolipin, and N-acetylglucosaminyl diradylglycerol were the major lipids present in most strains examined except for strain ATCC 10779, the parent of strain E88, the first C. tetani strain to have its genome sequenced.

Identification of undecaprenyl phosphate-beta-D-galactosamine in Francisella novicida and its function in lipid A modification.

Francisella tularensis is a highly infectious pathogen that causes tularemia. Francisella lipid A contains an unusual galactosamine (GalN) unit, attached to its 1-phosphate moiety. Two genes, flmF2 and flmK, are required for the addition of GalN to Francisella lipid A, but the relevant enzymes and the GalN donor substrate have not been characterized.

Structural analysis and detection of biological inositol pyrophosphates reveal that the family of VIP/diphosphoinositol pentakisphosphate kinases are 1/3-kinases.

We have characterized the positional specificity of the mammalian and yeast VIP/diphosphoinositol pentakisphosphate kinase (PPIP5K) family of inositol phosphate kinases. We deployed a microscale metal dye detection protocol coupled to a high performance liquid chromatography system that was calibrated with synthetic and biologically synthesized standards of inositol pyrophosphates. In addition, we have directly analyzed the structures of biological inositol pyrophosphates using two-dimensional 1H-1H and 1H-31P nuclear magnetic resonance spectroscopy.

Isolation and characterization of karlotoxin 1, a new amphipathic toxin from Karlodinium veneficum.

The karlotoxins (KmTxs) are a family of compounds produced by the dinoflagellate Karlodinium veneficum that cause membrane permeabilization. The structure of KmTx 1, determined using extensive 2D NMR spectroscopy, is very similar to the amphidinols and related compounds, though KmTx 1 features unique structural modifications of the conserved core region. The structure of KmTx 1 differs from that reported for KmTx 2, the only other reported karlotoxin to date, in lacking chlorination at its terminal alkene and possessing a hydrophobic arm that is two carbons longer.

Assessment of citrate concentrations in citrus fruit-based juices and beverages: implications for management of hypocitraturic nephrolithiasis.

Background And Purpose: Dietary intake of citrate in the form of citrus juices (eg, lemonade, orange juice) will enhance urinary citrate excretion, a valuable benefit for patients with hypocitraturic calcium oxalate nephrolithiasis. While information on citrate concentrations in select citrus juices is available, data on citrate concentrations of commercially available beverages (juice and otherwise) are limited. Using nuclear magnetic resonance spectroscopy (1H NMR), we report citrate concentrations of several beverages to help guide dietary recommendations aimed at increasing urinary citrate excretion and correcting hypocitraturia.

Biochemical analysis of inositol phosphate kinases.

Lipid-derived inositol phosphates (IPs) are a complex group of second messengers generated by the sequential phosphorylation of inositol 1,4,5-trisphosphate (IP(3)). Synthetic pathways leading from IP(3) to the formation of inositol tetrakisphosphate IP(4), inositol pentakisphosphate IP(5), inositol hexakisphosphate IP(6), and inositol pyrophosphates PP-IPs have been elucidated in eukaryotes from yeast to human. Studies have attributed a variety of cellular functions to IPs, highlighting the importance of understanding how the pathways for their synthesis are regulated.

A conserved family of enzymes that phosphorylate inositol hexakisphosphate.

Inositol pyrophosphates are a diverse group of high-energy signaling molecules whose cellular roles remain an active area of study. We report a previously uncharacterized class of inositol pyrophosphate synthase and find it is identical to yeast Vip1 and Asp1 proteins, regulators of actin-related protein-2/3 (ARP 2/3) complexes. Vip1 and Asp1 acted as enzymes that encode inositol hexakisphosphate (IP6) and inositol heptakisphosphate (IP7) kinase activities.

Attenuated virulence of a Francisella mutant lacking the lipid A 4'-phosphatase.

Francisella tularensis causes tularemia, a highly contagious disease of animals and humans, but the virulence features of F. tularensis are poorly defined. F.

Structure and biosynthesis of free lipid A molecules that replace lipopolysaccharide in Francisella tularensis subsp. novicida.

Francisella tularensis subsp. novicida U112 phospholipids, extracted without hydrolysis, consist mainly of phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine, and two lipid A species, designated A1 and A2. These lipid A species, present in a ratio of 7:1, comprise 15% of the total phospholipids, as judged by 32Pi labeling.

An outer membrane enzyme encoded by Salmonella typhimurium lpxR that removes the 3'-acyloxyacyl moiety of lipid A.

The Salmonella and related bacteria modify the structure of the lipid A portion of their lipopolysaccharide in response to environmental stimuli. Some lipid A modifications are required for virulence and resistance to cationic antimicrobial peptides. We now demonstrate that membranes of Salmonella typhimurium contain a novel hydrolase that removes the 3'-acyloxyacyl residue of lipid A in the presence of 5 mM Ca2+.

Kdo2-Lipid A of Escherichia coli, a defined endotoxin that activates macrophages via TLR-4.

The LIPID MAPS Consortium (www.lipidmaps.org) is developing comprehensive procedures for identifying all lipids of the macrophage, following activation by endotoxin.

A Leptospira interrogans enzyme with similarity to yeast Ste14p that methylates the 1-phosphate group of lipid A.

Distinct from other spirochetes, cells of Leptospira interrogans contain orthologues of all the Escherichia coli lpx genes required for lipid A biosynthesis, but they synthesize a modified form of lipopolysaccharide that supposedly activates toll-like receptor 2 (TLR2) instead of TLR4. The recent determination of the L. interrogans lipid A structure revealed an unprecedented O-methylation of its 1-phosphate group (Que-Gewirth, N.

A formyltransferase required for polymyxin resistance in Escherichia coli and the modification of lipid A with 4-Amino-4-deoxy-L-arabinose. Identification and function oF UDP-4-deoxy-4-formamido-L-arabinose.

Modification of the phosphate groups of lipid A with 4-amino-4-deoxy-L-arabinose (L-Ara4N) is required for resistance to polymyxin and cationic antimicrobial peptides in Escherichia coli and Salmonella typhimurium. We previously demonstrated that the enzyme ArnA catalyzes the NAD+-dependent oxidative decarboxylation of UDP-glucuronic acid to yield the UDP-4''-ketopentose, uridine 5'-diphospho-beta-(L-threo-pentapyranosyl-4''-ulose), which is converted by ArnB to UDP-beta-(L-Ara4N). E.

Oxidation and transamination of the 3"-position of UDP-N-acetylglucosamine by enzymes from Acidithiobacillus ferrooxidans. Role in the formation of lipid a molecules with four amide-linked acyl chains.

Lipid A, a major component of the outer membranes of Escherichia coli and other Gram-negative bacteria, is usually constructed around a beta-1',6-linked glucosamine disaccharide backbone. However, in organisms like Acidithiobacillus ferrooxidans, Leptospira interrogans, Mesorhizobium loti, and Legionella pneumophila, one or both glucosamine residues are replaced with the sugar 2,3-diamino-2,3-dideoxy-d-glucopyranose. We now report the identification of two proteins, designated GnnA and GnnB, involved in the formation of the 2,3-diamino-2,3-dideoxy-d-glucopyranose moiety.

A methylated phosphate group and four amide-linked acyl chains in leptospira interrogans lipid A. The membrane anchor of an unusual lipopolysaccharide that activates TLR2.

Leptospira interrogans differs from other spirochetes in that it contains homologs of all the Escherichia coli lpx genes required for the biosynthesis of the lipid A anchor of lipopolysaccharide (LPS). LPS from L. interrogans cells is unusual in that it activates TLR2 rather than TLR4.

Dendritic molecular capsules for hydrophobic compounds.

Reichardt's dye, a highly solvatochromic dye, was encapsulated within poly (glycerol succinic acid) ([Gn]-PGLSA-OH) dendrimers to investigate the interior environment of these dendritic macromolecules. The absorption maximum for the encapsulated Reichardt's dye in water was indicative of a relatively high dielectric constant present within the dye/dendrimer complex. (1)H NMR of the encapsulated complex showed the presence of aromatic protons from Reichardt's dye along with the aliphatic protons of the dendrimer.

Origin of lipid A species modified with 4-amino-4-deoxy-L-arabinose in polymyxin-resistant mutants of Escherichia coli. An aminotransferase (ArnB) that generates UDP-4-deoxyl-L-arabinose.

In Escherichia coli and Salmonella typhimurium, addition of the 4-amino-4-deoxy-l-arabinose (l-Ara4N) moiety to the phosphate group(s) of lipid A is required for resistance to polymyxin and cationic antimicrobial peptides. We have proposed previously (Breazeale, S. D.

A mannosyl transferase required for lipopolysaccharide inner core assembly in Rhizobium leguminosarum. Purification, substrate specificity, and expression in Salmonella waaC mutants.

The lipopolysaccharide (LPS) core domain of Gram-negative bacteria plays an important role in outer membrane stability and host interactions. Little is known about the biochemical properties of the glycosyltransferases that assemble the LPS core. We now report the purification and characterization of the Rhizobium leguminosarum mannosyl transferase LpcC, which adds a mannose unit to the inner 3-deoxy-d-manno-octulosonic acid (Kdo) moiety of the LPS precursor, Kdo(2)-lipid IV(A).

Synthesis and characterization of carbohydrate-based phospholipids.

Novel carbohydrate-based phospholipids containing two saturated C(12) (dilauroyl ribo-phosphocholine) (DLRPC), C(14) (dimyristoyl ribo-phosphocholine) (DMRPC), and C(20) (diarachadonyl ribo-phosphocholine) (DARPC) carboxylic acid chains were synthesized. The physical properties of the supramolecular structures formed by these compounds were compared to those formed by their direct glycerol analogues dilauroyl phosphocholine (DLPC), dimyristoyl phosphocholine (DMPC), and diarachadonyl phosphocholine (DAPC). Modulated differential scanning calorimetry (MDSC) and X-ray diffraction data indicated that with chain lengths < or =14 carbons, the carbohydrate backbone increased the thermal stability of the bilayer below the phase-transition temperature (T(m)) as compared to the glycerol-based lipids.

The Escherichia coli gene encoding the UDP-2,3-diacylglucosamine pyrophosphatase of lipid A biosynthesis.

UDP-2,3-diacylglucosamine hydrolase is believed to catalyze the fourth step of lipid A biosynthesis in Escherichia coli. This reaction involves pyrophosphate bond hydrolysis of the precursor UDP-2,3-diacylglucosamine to yield 2,3-diacylglucosamine 1-phosphate and UMP. To identify the gene encoding this hydrolase, E.

Oxidative decarboxylation of UDP-glucuronic acid in extracts of polymyxin-resistant Escherichia coli. Origin of lipid a species modified with 4-amino-4-deoxy-L-arabinose.

Addition of the 4-amino-4-deoxy-l-arabinose (l-Ara4N) moiety to the phosphate groups of lipid A is implicated in bacterial resistance to polymyxin and cationic antimicrobial peptides of the innate immune system. The sequences of the products of the Salmonella typhimurium pmrE and pmrF loci, both of which are required for polymyxin resistance, recently led us to propose a pathway for l-Ara4N biosynthesis from UDP-glucuronic acid (Zhou, Z., Lin, S.

Application of a Radical Methodology toward the Synthesis of d,l-5alpha-Pregnanes and Related Steroids: A Stereoselective Radical Cascade Approach.

A stereoselective radical cascade cyclization to 5alpha-pregnanes is presented. Oxidative free radical cyclization of an appropriately substituted chloro cyano ester polyene was used to introduce the all trans stereochemistry in the steroid nucleus. The cyano group was utilized to introduce a C-8beta angular hydrogen, while the chloro ester moiety served as an entry to the geminal hydrogens at C-4.