Uptake hydrogenase in fast-growing strains of Rhizobium sp. (Sesbania) in relation to nitrogen fixation

Eight of 104 fast-growing strains of Rhizobium sp. ( Sesbania ) tested possessed a hydrogen recycling system. Depression of uptake hydrogenase occurred in poor as well as in rich carbon media. A statistically significant increase (> 22%) in total plant nitrogen content and dry matter yield was observed by the inoculation of hup + strains over hup ^- strains.     

Transfer of Brassica tournefartii (TT) genes to allotetraploid oilseed Brassica species (B. juncea AABB, B. napus AACC, B. carinata BBCC): homoeologous pairing is more pronounced in the three-genome hybrids (TACC, TBAA, TCAA, TCBB) as compared to allodiploids (TA, TB, TC)

For the transfer of genes from B. tournefortii (TT) to the allotetraploid oilseed brassicas, B. juncea AABB, B. carinata BBCC and B. napus AACC, B. tournefortii was first crossed with the three basic diploid species, B. campestris (AA), B. nigra (BE) and B. oleracea (CC), to produce the allodiploids TA, TB and TC. These were tetraploidized by colchicine treatment to produce the allotetraploids TTAA, TTBB and TTCC, which were further crossed with B. juncea and B. napus to produce three-genome hybrids with substitution-type genomic configurations: TACC, TBAA and TCAA. These hybrids along with another hybrid TCBB produced earlier, the three allodiploids, their allotetraploids and the four diploid parent species were studied for their male meiotic behaviour. The diploid parent and the allotetraploids (TTAA, TTBB and TTCC) showed regular meiosis although the pollen viability was generally low in the allotetraploids. In the allodiploids (TA, TB and TC) only some end-to-end associations were observed without any clearly discernible chiasmata or exchange points. Chromosomes involved in end-to-end associations were randomly distributed at the metaphase/anaphase-I stages. In contrast, the three-genome hybrids (TACC, TBAA, TCAA and TCBB) showed normal bivalents whose number exceeded the expected bivalent values. Bivalents arising out of homoeologous pairing were indistinguishable from normal pairs by their disjunction pattern but could be distinguished on the basis of the heteromorphy of the homoeologous chromosomes. The three-genome hybrids could be backcrossed to allotetraploid oilseed brassicas as they had some fertility. In contrast, the allodiploids could neither be selfed nor back-crossed. On the basis of their meiotic stability, in terms of more pronounced homoeologous pairing and fertility for backcrossing, the three-genome configurations provide the best possible situation for the introgression of alien genes from the secondary gene pool to the allotetraploid oilseed crops B. juncea, B. napus and B. carinata.     

Effect of carbon dioxide concentration on the bioleaching of a pyrite-arsenopyrite ore concentrate

The effect of carbon dioxide concentration on the bacterial leaching of a pyrite-arsenopyrite ore concentrate was studied in continuous-flow reactors. Steady-state operation with two feed slurry densities, 6 wt% and 16 wt% solids, were tested for the effect of carbon dioxide concentration. Bacterial growth rates were estimated via the measurement of carbon dioxide consumption rates. Aqueous-phase carbon dioxide concentrations in excess of 10 mg/L were found to be inhibitory to bacterial growth. (c) 1993 John Wiley & Sons, Inc.     

The use of s-2-cyanoethyl phosphorothioate in the preparation of oligo 5''-deoxy-5''-thiothymidylates.

An improvement of our strategy for the stepwise synthesis of oligo 5'-deoxy-5'-thiodeoxyribonucleotides [Chladek and Nagyvary (1972) J. Amer. Chem. Soc. 94, 2079] involves the use of 5'-O-tosylthymidine 3'-S-2-cyanoethyl phosphorothioate. The displacement of the tosylate by thymidine 3'-phosphorothioate and subsequent alkaline deblocking afforded the dinucleotide (Tps)2. The process of displacement and deblocking was repeated three more times at an average yield of 30 percent per step. The corresponding bifunctional derivative of deoxyadenosine was found much less reactive and practically unsuitable for repeated chain elongation. The ORD and CD spectra of the analogs are similar to those of the natural oligonucleotides.     

Inhibition of D-glucose uptake by isatin in rat intestine: effect of harmaline and various sulfhydryl reagents

The effect of isatin (indole-2,3-dione) on D-glucose uptake has been studied in rat intestine. Isatin at 6 mM concentration significantly inhibited both the sugar uptake and transmural (mucosal to serosal side) transport in the intestine. The suppression of glucose uptake by isatin was irreversible. Similar to the action of various SH-group-reacting agents, isatin inhibited the sugar uptake, presumably by binding to membrane sulfhydryl groups through a covalent linkage. Isatin-induced reduction in glucose uptake was unaffected by pH (between 5.5 and 8.4) and by DTT addition to incubation medium. Inhibition of sugar uptake by isatin and harmaline was additive in nature; this suggested that these compounds interact at different sites on the microvillus membrane surface.     

Vikodak - A Modular Framework for Inferring Functional Potential of Microbial Communities from 16S Metagenomic Datasets

Background

The overall metabolic/functional potential of any given environmental niche is a function of the sum total of genes/proteins/enzymes that are encoded and expressed by various interacting microbes residing in that niche. Consequently, prior (collated) information pertaining to genes, enzymes encoded by the resident microbes can aid in indirectly (re)constructing/ inferring the metabolic/ functional potential of a given microbial community (given its taxonomic abundance profile). In this study, we present Vikodak—a multi-modular package that is based on the above assumption and automates inferring and/ or comparing the functional characteristics of an environment using taxonomic abundance generated from one or more environmental sample datasets. With the underlying assumptions of co-metabolism and independent contributions of different microbes in a community, a concerted effort has been made to accommodate microbial co-existence patterns in various modules incorporated in Vikodak.

Results

Validation experiments on over 1400 metagenomic samples have confirmed the utility of Vikodak in (a) deciphering enzyme abundance profiles of any KEGG metabolic pathway, (b) functional resolution of distinct metagenomic environments, (c) inferring patterns of functional interaction between resident microbes, and (d) automating statistical comparison of functional features of studied microbiomes. Novel features incorporated in Vikodak also facilitate automatic removal of false positives and spurious functional predictions.

Conclusions

With novel provisions for comprehensive functional analysis, inclusion of microbial co-existence pattern based algorithms, automated inter-environment comparisons; in-depth analysis of individual metabolic pathways and greater flexibilities at the user end, Vikodak is expected to be an important value addition to the family of existing tools for 16S based function prediction.

Availability and Implementation

A web implementation of Vikodak can be publicly accessed at: http://metagenomics.atc.tcs.com/vikodak. This web service is freely available for all categories of users (academic as well as commercial).     

Ligand modulates VDR-Ser/Thr protein phosphatase interaction and p70S6 kinase phosphorylation in a cell-context-dependent manner

We have recently shown that in colon cancer cells, Vitamin D receptor (VDR) interacts with the catalytic subunit of Ser/Thr protein phosphatases, PP1c and PP2Ac, and induces their enzymatic activity in a ligand-dependent manner. The VDR-PP1c and VDR-PP2Ac interactions were ligand independent in vivo, and 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3))-mediated increase in VDR-associated phosphatase activity resulted in dephosphorylation and inactivation of p70S6 kinase in colon cancer cells. Here, we demonstrate that in myeloid leukemia cells, 1,25(OH)(2)D(3) treatment increased the Thr389 phosphorylation of p70S6 kinase. Accordingly, 1,25(OH)(2)D(3) decreased VDR-associated Ser/Thr protein phosphatase activity by dissociating VDR-PP1c and VDR-PP2Ac interactions. Further, 1,25(OH)(2)D(3) increased the association between VDR and Thr389 phosphorylated p70S6 kinase. Finally, by using non-secosteroidal VDR ligands, we demonstrate a separation between transactivation and p70S6 kinase phosphorylation activities of VDR and show pharmacologically that p70S6 kinase phosphorylation correlates with HL-60 cell differentiation.     

Indene bioconversion by a toluene inducible dioxygenase of Rhodococcus sp. I24

Rhodococcus sp. I24 can oxygenate indene via at least three independent enzyme activities: (i) a naphthalene inducible monooxygenase (ii) a naphthalene inducible dioxygenase, and (iii) a toluene inducible dioxygenase (TID). Pulsed field gel analysis revealed that the I24 strain harbors two megaplasmids of 340 and 50 kb. Rhodococcus sp. KY1, a derivative of the I24 strain, lacks the 340 kb element as well as the TID activity. Southern blotting and sequence analysis of an indigogenic, I24-derived cosmid suggested that an operon encoding a TID resides on the 340 kb element. Expression of the tid operon was induced by toluene but not by naphthalene. In contrast, naphthalene did induce expression of the nid operon, encoding the naphthalene dioxygenase in I24. Cell free protein extracts of Escherichia coli cells expressing tidABCD were used in HPLC-based enzyme assays to characterize the indene bioconversion of TID in vitro. In addition to 1-indenol, indene was transformed to cis-indandiol with an enantiomeric excess of 45.2% of cis-(1S,2R)-indandiol over cis-(1R,2S)-indandiol, as revealed by chiral HPLC analysis. The K_m of TID for indene was 380 M. The enzyme also dioxygenated naphthalene to cis-dihydronaphthalenediol with an activity of 78% compared to the formation of cis-indandiol from indene. The K_m of TID for naphthalene was 28 M. TID converted only trace amounts of toluene to 1,2-dihydro-3-methylcatechol after prolonged incubation time. The results indicate the role of the tid operon in the bioconversion of indene to 1-indenol and cis-(1S,2R)-indandiol by Rhodococcus sp. I24.     

Plasticity in structure and interactions is critical for the action of indolicidin,an antibacterial peptide of innate immune origin

The comparative analysis of two cationic antibacterial peptides of the cathelicidin family-indolicidin and tritrypticin-enabled addressing the structural features critical for the mechanism of indolicidin activity. Functional behavior of retro-indolicidin was found to be identical to that of native indolicidin. It is apparent that the gross conformational propensities associated with retro-peptides resemble those of the native sequences, suggesting that native and retro-peptides can have similar structures. Both the native and the retro-indolicidin show identical affinities while binding to endotoxin, the initial event associated with the antibacterial activity of cationic peptide antibiotics. The indolicidin-endotoxin binding was modeled by docking the indolicidin molecule in the endotoxin structure. The conformational flexibility associated with the indolicidin residues, as well as that of the fatty acid chains of endotoxin combined with the relatively strong structural interactions, such as ionic and hydrophobic, provide the basis for the endotoxin-peptide recognition. Thus, the key feature of the recognition between the cationic antibacterial peptides and endotoxin is the plasticity of molecular interactions, which may have been designed for the purpose of maintaining activity against a broad range of organisms, a hallmark of primitive host defense.     

Restriction fragment length polymorphism of rRNA operons for discrimination and intergenic spacer sequences for cataloging of Bacillus subtilis sub-groups

Restriction fragment length polymorphism of rRNA operons (RFLP) and 16S-23S rRNA intergenic region (ISR) sequences of Bacillus subtilis subsp. subtilis, B. subtilis subsp. spizizenii, and B. atrophaeus were compared. ISR sequences of the B. subtilis subspecies were extremely similar (W23 versus 168 rrn H, J, G,W; 96.8%; rrn D, E; 98.4%; rrnB; 97.9%) and, therefore, not useful for their differentiation. However, RFLP of rRNA operons of the B. subtilis subspecies were distinct in terms of numbers and organization within the genome (e.g. the 168 sub-group generally contained 8.3- and 8.0-kb fragments absent in the W23 sub-group). The more distantly related B. atrophaeus was distinct from both B. subtilis subspecies in terms of ISR sequence and rRNA operon number and organization. RFLP of rRNA operons discriminates the two sub-groups of Bacillus subtilis that are indistinguishable by ISR sequence. However, ISR sequence defines the relatedness of B. subtilis to other species (e.g. B. atrophaeus) within the genus Bacillus.