Shift from widespread symbiont infection of host tissues to specific colonization of gills in juvenile deep-sea mussels

The deep-sea mussel Bathymodiolus harbors chemosynthetic bacteria in its gills that provide it with nutrition. Symbiont colonization is assumed to occur in early life stages by uptake from the environment, but little is known about this process. In this study, we used fluorescence in situ hybridization to examine symbiont distribution and the specificity of the infection process in juvenile B. azoricus and B. puteoserpentis (4–21 mm). In the smallest juveniles, we observed symbionts, but no other bacteria, in a wide range of epithelial tissues. This suggests that despite the widespread distribution of symbionts in many different juvenile organs, the infection process is highly specific and limited to the symbiotic bacteria. Juveniles⩾9 mm only had symbionts in their gills, indicating an ontogenetic shift in symbiont colonization from indiscriminate infection of almost all epithelia in early life stages to spatially restricted colonization of gills in later developmental stages.     

A novel imidazole nucleoside containing a diaminodihydro-S-triazine as a substituent: inhibitory activity against the West Nile virus NTPase/helicase

The attempted synthesis of a ring-expanded guanosine (1) containing the imidazo[4,5-e][1,3]diazepine ring system by condensation of 1-(2'-deoxy-beta-D-erythropentofuranosyl)-4-ethoxycarbonylimidazole-5-carbaldehyde (2) with guanidine resulted in the formation of an unexpected product, 1-(2'-deoxy-beta-D-erythropentofuranosyl)-5-(2, 4-diamino-3, 6-dihydro-1,3, 5-triazin-6-yl)imidazole-4-carboxamide (7). The structure as well as the pathway of formation of 7 was corroborated by isolation of the intermediate, followed by its conversion to the product. Nucleoside 7 showed promising in vitro anti-helicase activity against the West Nile virus NTPase/helicase with an IC50 of 3-10 microg/mL.     

An analogue of AICAR with dual inhibitory activity against WNV and HCV NTPase/helicase: synthesis and in vitro screening of 4-carbamoyl-5-(4,6-diamino-2,5-dihydro-1,3,5-triazin-2-yl)imidazole-1-beta-D-ribofuranoside

The title compound (4) was synthesized by the reaction of ethyl 1-(2,3,5-tri-O-benzoyl-beta-d-ribofuranosyl)-5-formylimidazole-4-carboxylate with excess guanidine in ethanol at reflux. Compound 4 was evaluated in vitro against NTPases/helicases of four different viruses of the Flaviviridae family, including the West Nile virus (WNV), hepatitis C virus (HCV), dengue virus (DENV), and the Japanese encephalitis virus (JEV), employing both an RNA and a DNA substrate. The compound showed activity against NTPase/helicase of WNV and HCV with an IC(50) of 23 and 37 microM, respectively, when a DNA substrate was employed, while no activity was observed when an RNA substrate was used. There was no activity against the NTPase/helicase of either DENV or JEV irrespective of whether an RNA or a DNA substrate was employed. Considering that Flaviviridae are RNA viruses, the observed absence of activity against an RNA substrate, but the presence of activity against a DNA substrate is intriguing and somewhat surprising. The preliminary studies show that compound 4 does not form a tight complex with either an RNA or a DNA substrate, suggesting that its mechanism of action may involve direct interaction with the enzyme.     

Influence of a lipid bilayer on the conformational behavior of amphotericin B derivatives — A molecular dynamics study

Amphotericin B (AmB) is an effective but very toxic antifungal antibiotic. In our laboratory a series of AmB derivatives of improved selectivity of action was synthesized and tested. To understand molecular basis of this improvement, comparative conformational studies of amphotericin B and its two more selective derivatives were carried out in an aqueous solution and in a lipid membrane. These molecular simulation studies revealed that within a membrane environment the conformational behavior of the derivatives differs significantly from the one observed for the parent molecule. Possible reasons for such a difference are analyzed. Furthermore, we hypothesize that the observed conformational transition within the polar head of AmB derivatives may lead to destabilization of antibiotic-induced transmembrane channels. Consequently, the selective toxicity of the derivatives should increase as ergosterol-rich liquid-ordered domains are more rigid and conformationally ordered than their cholesterol-containing counterparts, and as such may better support less stable channel structure.     

Mechanism for cyclization reaction by clavaminic acid synthase. Insights from modeling studies

The mechanism of the oxidative cyclization reaction catalyzed by clavaminic acid synthase (CAS) was studied in silico. First, a classical molecular dynamics (MD) simulation was performed to obtain a realistic structure of the CAS-Fe(IV)=O-succinate-substrate complex; then potential of mean force (PMF) was calculated to assess the feasibility of the beta-lactam ring, more specifically its C4' corner, approaching the oxo atom. Based on the MD structure, a relatively large model of the active site region was selected and used in the B3LYP investigation of the reaction mechanism. The computational results suggest that once the oxoferryl species is formed, the oxidative cyclization catalyzed by CAS most likely involves either a mechanism involving C4'(S)-H bond cleavage of the monocyclic beta-lactam ring, or a biosynthetically unprecedented mechanism comprising (1) oxidation of the hydroxyl group of PCA to an O-radical, (2) retro-aldol-like decomposition of the O-radical to an aldehyde and a C-centered radical, which is stabilized by the captodative effect, (3) abstraction of a hydrogen atom from the C4'(S) position of the C-centered radical by the Fe(III)-OH species yielding an azomethine ylide, and (4) 1,3-dipolar cycloaddition to the ylide with aldehyde acting as a dipolarophile. Precedent for the new proposed mechanism comes from the reported synthesis of oxapenams via 1,3-dipolar cycloaddition reactions of aldehydes and ketones.     

Effects of Tomatex /Triacontanol/ on chlorophyll fluorescence and tomato (Lycopersicon esculentum Mill.) yields

The influence of triacontanol in a form of Tomatex preparation on basic indices of chlorophyll fluorescence in tomato leaves (Delfina cv.), yield of fruits, and dry matter content in fruits was evaluated in a pot experiment situated in vegetation hall in 1999. Tomatex was applied into roots at seedling stage (6–7 leaves) or at the stage of seedling and flowering of the 2^nd inflorescence bunch. Plants were given by 0.3, 3.0, and 30 μg triacontanol per pot at a single dosage. Results obtained have shown that triacontanol regardless of the dose applied, significantly increased the maximal efficiency of PSII photochemistry in the dark (F_v/Fm), the efficiency of excitation capture by open PSII reaction centers (F_v’/F_m’), the actual quantum yield of PSII electron transport in the light-adapted state (Φ_PSII), the photochemical quenching coefficient (Q_p). However, nonphotochemical quenching coefficient (Q_n) and non-radiative dissipation (NPQ) were decreased. Plants treated with triacontanol at the doses of 0.3 and 3.0 μg had significantly higher yields of fruits than control. No differences were found between plants treated once and twice with the growth regulator. Triacontanol did not show univocal effects on dry matter content in fruits either.     

Bacteriolytic effect of cessation of glucosamine supply, induced by specific inhibition of glucosamine-6-phosphate synthetase

The antibiotic tetaine (bacilysin) and its C-terminal epoxyaminoacid--anticapsin--are powerful inhibitors of glucosamine-6-phosphate synthetase (EC 5.3.1.19.) in cell-free extracts of Escherichia coli K-12. Tetaine acts on growing cells as a bactericidal agent. This bactericidal action, measured from 10 to 160 muM concentration, is a consequence of the induction of lysis of growing cells. The induction of lysis by tetaine is compared with the lytic action of some beta-lactams. Hypertonic medium, destruction of the antibiotic, presence of chloramphenicol or the addition of N-acetylglucosamine protect E. coli K-12 cells against lysis induced by tetaine. These effects are compared with those observed in the presence of penicillin G. The results indicate that inhibition of early or late stages of peptidoglycan synthesis all result in more or less the same consequence, i.e. death via cell lysis.