RCAI-17, 22, 24-26, 29, 31, 34-36, 38-40, and 88, the analogs of KRN7000 with a sulfonamide linkage: their synthesis and bioactivity for mouse natural killer T cells to produce Th2-biased cytokines

RCAI-17, 22, 24-26, 29, 31, 34-36, 38-40, and 88, the analogs of KRN7000 with a sulfonamide linkage instead of an amide bond, were synthesized to examine their bioactivity for mouse natural killer (NK) T cells. RCAI-17, 22, 24-26, 29, 31, 34-36, and 88 are the aromatic sulfonamide analogs, while RCAI-39 and 40 are the aliphatic ones. RCAI-38 is a C-galactoside analog of RCAI-26, which is the p-toluenesulfonamide analog of KRN7000. According to their bioassay, these sulfonamide analogs were shown to be the stimulants of mouse NKT cells to induce the production of Th2-biased cytokines in vitro, while RCAI-38 did not induce any cytokine production.     

Purification and characterization of cyclic maltosyl-(1-->6)-maltose hydrolase and alpha-glucosidase from an Arthrobacter globiformis strain

Cyclic maltosyl-maltose [CMM, cyclo-[-->6)-alpha-D-Glcp-(1-->4)-alpha-D-Glcp-(1-->6)-alpha-D-Glcp-(1-->4)-alpha-D-Glcp-(1-->]], a novel cyclic tetrasaccharide, has a unique structure. Its four glucose residues are joined by alternate alpha-1,4 and alpha-1,6 linkages. CMM is synthesized from starch by the action of 6-alpha-maltosyltransferase from Arthrobacter globiformis M6. Recently, we determined the mechanism of extracellular synthesis of CMM, but the degrading pathway of the saccharide remains unknown. Hence we tried to identify the enzymes involved in the degradation of CMM to glucose from the cell-free extract of the strain, and identified CMM hydrolase (CMMase) and alpha-glucosidase as the responsible enzymes. The molecular mass of CMMase was determined to be 48.6 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and 136 kDa by gel filtration column chromatography. The optimal pH and temperature for CMMase activity were 6.5 and 30 degrees C. The enzyme remained stable from pH 5.5 to 8.0 and up to 25 degrees C. CMMase hydrolyzed CMM to maltose via maltosyl-maltose as intermediates, but it did not hydrolyze CMM to glucose, suggesting that it is a novel hydrolase that hydrolyzes the alpha-1,6-linkage of CMM. The molecular mass of alpha-glucosidase was determined to be 60.1 kDa by SDS-PAGE and 69.5 kDa by gel filtration column chromatography. The optimal pH and temperature for alpha-glucosidase activity were 7.0 and 35 degrees C. The enzyme remained stable from pH 7.0 to 9.5 and up to 35 degrees C. alpha-Glucosidase degraded maltosyl-maltose to glucose via panose and maltose as intermediates, but it did not degrade CMM. Furthermore, when CMMase and alpha-glucosidase existed simultaneously in a reaction mixture containing CMM, glucose was detected as the final product. It was found that CMM was degraded to glucose by the synergistic action of CMMase and alpha-glucosidase.     

Chemical communication in mating behaviour of the slave-making ant <Emphasis Type="Italic">Polyergus rufescens</Emphasis> (Hymenoptera,Formicidae): 3-ethyl-4-methylpentanol as a critical component of the queen sex pheromone

The aim of the research reported here was to determine whether 3-ethyl-4-methylpentanol, a minor but crucial component of the sex pheromone of the North American slave-making ant species Polyergus breviceps, was also a component of the sex pheromone of the European congener Polyergus rufescens. Thus, the contents of mandibular glands of P. rufescens virgin queen were extracted and analysed. The main component of the extracts was methyl 6-methylsalicylate and 3-ethyl-4-methylpentanol was identified as one of several minor components. Further analyses showed that the insects produce mainly the (R)-enantiomer of the alcohol. Males’ responses to various blends of methyl 6-methylsalicylate with the racemate or the pure enantiomers of 3-ethyl-4- methylpentanol were tested in field behavioural bioassays. The data showed that blends of methyl 6- methylsalicylate and 3-ethyl-4-methylpentanol were strongly synergistic, with the most active ratios being biased toward the first component. The addition of other minor components to the binary blend neither increased nor decreased responses by males. Only the (R)-enantiomer of the alcohol was biologically active; its antipode did not inhibit attraction. The results are discussed in terms of the evolution of signals, and are compared with the results previously obtained for the allopatric species Polyergus breviceps. Received 12 November 2007; revised 10 January 2008; accepted 11 January 2008.     

The outer membrane TolC is involved in cysteine tolerance and overproduction in <Emphasis Type="Italic">Escherichia coli</Emphasis>

l-Cysteine is an important amino acid in terms of its industrial applications. We previously found marked production of l-cysteine directly from glucose in recombinant Escherichia coli cells by the combination of enhancing biosynthetic activity and weakening the degradation pathway. Further improvements in l-cysteine production are expected to use the amino acid efflux system. Here, we identified a novel gene involved in l-cysteine export using a systematic and comprehensive collection of gene-disrupted E. coli K-12 mutants (the Keio collection). Among the 3,985 nonessential gene mutants, tolC-disrupted cells showed hypersensitivity to l-cysteine relative to wild-type cells. Gene expression analysis revealed that the tolC gene encoding the outer membrane channel is essential for l-cysteine tolerance in E. coli cells. However, l-cysteine tolerance is not mediated by TolC-dependent drug efflux systems such as AcrA and AcrB. It also appears that other outer membrane porins including OmpA and OmpF do not participate in TolC-dependent l-cysteine tolerance. When a low-copy-number plasmid carrying the tolC gene was introduced into E. coli cells with enhanced biosynthesis, weakened degradation, and improved export of l-cysteine, the transformants exhibited more l-cysteine tolerance and production than cells carrying the vector only. We concluded that TolC plays an important role in l-cysteine tolerance probably due to its export ability and that TolC overexpression is effective for l-cysteine production in E. coli. Natthawut Wiriyathanawudhiwong and Iwao Ohtsu contributed equally to this work.     

Synthesis,cleavage, and antifungal activity of a number of novel,water-soluble ester prodrugs of antifungal triazole CS-758

In this study, the synthesis and evaluation of a number of esters of CS-758 as injectable prodrugs are described. Phosphoryl ester 1a was soluble in water (>30 mg/mL) and was converted to CS-758 in human liver microsome. It was also converted to CS-758 in rats after iv administration, wherein the bioavailability of CS-758 was 53%. Compound 1a (iv) reduced the viable cell counts in kidneys in a murine systemic Candida albicans infection model, wherein the effect was comparable to or slightly superior to that of CS-758 (po). The prodrug 1a proved to be a promising injectable antifungal agent whose further evaluation is warranted.