An in-silico investigation of fluoride ions impact on pancreatic lipase.

Fluorine is a halogen beneficial to teeth and bones at a lower concentration. But in excess, it is a toxin and causes adverse effects. Fluoride is toxic to enzymes generally when it inhibits the enzyme activity involved in metabolic pathways. Here we study invitro and invivo findings on the interaction of fluoride on the enzymes Aconitase, Adenylyl cyclase, Arginase, Cytochrome-c-oxidase, Glucose-6-phosphatase, Protein phosphatase, Succinate dehydrogenase from liver and lipase from pancreas by using molecular docking and simulations to gain insights into the mechanism by which fluoride modifies the activity of pancreatic lipase. our molecular modeling and docking studies identified that lipase is the most strongly inhibited enzyme compared to other enzymes mentioned above with -0.42 Kcal/mol binding energy and 495.78 milli molar of predicted IC50 value with interaction with Phe227 residue. To further validate this, we have taken the lipase enzyme in presence of fluoride ions for molecular dynamic simulations of 100 ns. To analyze the impact of fluoride ions on the lipase dynamics, two different simulations of 100 ns each were performed. In one simulation, we have simulated lipase in its apo form in the aqueous environment without any fluoride ions and in another simulation lipase in its apo form was kept in the presence of randomly placed fluoride ions countered with sodium ions to maintain the pH as neutral. The simulation analysis revealed that major fluctuations in lipase was observed between 230 and 300 residues in presence of fluoride ions. Interestingly, this is the exact location of the "lid" like acting loop of residues responsible for the inward/outward movement of the substrate to lipase catalytically active site containing catalytic triad of residues Leu153, His263, and Pro177. His263 residue random flip is believed to be the critical incident that causes the substrate's inward/outward movement at the catalytically active site coordinated by "lid" opening, providing enough space for the substrate.

Microsecond simulation analysis of carbonic anhydrase - II in complex with (+)-cathechin revealed molecular interactions responsible for its amelioration effect on fluoride toxicity.

Fluorosis is a chronic condition caused by overexposure to fluoride, marked by impaired dental, skeletal, and non-skeletal health. In presence of excess fluoride ions, in severe cases calcification of the ligaments observed. Earlier studies have suggested that the disruption of carbonic anhydrase activity via ionic homeostasis change was associated with F toxicity. In a recent study, it was demonstrated that Tamarind fruit extract was effective in increasing the urinary F excretion in male Wistar rats via studying the mRNA expression of carbonic anhydrase II (CA II) in kidney homogenates using western blotting, immunohistochemistry and quantitative Realtime PCR based studies. We have carried out this study to understand the detailed molecular level interactions responsible for this tamarind extract based (+)-cathechin compound towards lowering the F toxicity via targeting CA-II. From our study, it was revealed that due to the ability of (+)-cathechin compound to bind tightly filling complete available space at the catalytically important site forming metal coordinated ionic bonds with His94, His96 and His119 residues helps in restricting F ions to interact with Zn ion located at the core of catalytic site responsible for its functionality. On the other hand, interaction of (+)-cathechin compound with Gln92 was observed to be critically important towards inducing conformational changes in CA-II, thus allowing (+)-cathechin compound to burry even deeply inside the catalytic site.Communicated by Ramaswamy H. Sarma.

Role of Thr199 residue in human ß-carbonic anhydrase-II pH-dependent activity elucidated by microsecond simulation analysis.

Carbonic anhydrases catalyze the reversible hydration of carbon dioxide to form bicarbonate, a reaction required for many functions such as carbon assimilation, pH acid-base homeostasis, respiration and photosynthesis via a zinc-hydroxide mechanism for carbon dioxide hydration. In earlier studies, it was revealed that Carbonic anhydrases are inactive at pH 7.5 and active at pH 8.4. This steep pH dependence for its activity led us to design this work to understand its mode of action at atomic level detail. In our microsecond simulation based analysis, it was revealed that the interaction between Glu106 and Thr199 plays a critically important role in its activity. Thr199 co-ordinated loop movement was observed to be acting as a lid, with 'open' and 'close' mechanism for substrate entry to the core of the catalytic site, where Zn-ion resides and executes its carbon dioxide hydration mechanism. On the other hand, decline in the total secondary structural elements percentage in the protein was observed in correspondence to the pH condition change. The α-helices between Thr125-Gly145 and Val150-Lys170 residues were especially noticed to be losing their structural integrity responsible for formation of dimer and tetramers. In conclusion, our analysis showed that the interaction between Glu106 and Thr199 is crucial for maintaining the structural integrity of the Thr199 coordinated loop, responsible for allowing substrate towards the catalytic site.Communicated by Ramaswamy H. Sarma.

Data on host specificity and symbiotic association between indigenous Rhizobium BD1 strain and Vigna radiata (green gram).

The main objective of this study is to use bio-inoculants in relative to specific legume plant diversity for, enhanced nodulation and plant growth. Method involves organically based selection of 36 rhizobial strains, of which 6 strains were isolated to assess the efficiency of relative host-specific inoculation on nodulation and development in legumes viz. Vigna radiata. All promising combinations of the preferred rhizobial strain inoculants were tested under sterile conditions for improving nodulation and to screen the best isolate to be evaluated for its enhanced characteristics through inoculation by field trial in various soils. It was observed that the strains from Bhadrachalam forest BD1 are highly host specific for Vigna radiata plants and when inoculated, improved nodulation and enhanced plant growth. Because of the novel characters in BD1, further studies were carried out and was identified as Rhizobium sp. BD1 (NCBI Accession no. MT577595). The percentage of nitrogen content in Vigna radiata ranged between 1.2% to 2.9%. This Rhizobium sp. BD1 was tested for the unraveling and amelioration of crop production in barren, polluted and agricultural soils which showed enhanced characteristics in Vigna radiata plants. This method may be employed across the globe of same climatic conditions for the retrieval of plants in soils that carry agriculture unsuccessfully.

Data on fluoride concentration profile in groundwater of rural habitats in Mahabubnagar district, Telangana, India.

Fluoride is an undesirable chemical in drinking water that can have dreadful concerns on health. The objective of this study is to establish the concentration profile of fluoride in groundwater used as drinking water in 31 rural habitats in Mahabubnagar district, Telangana State, India. Drinking water samples from bore wells/hand pumps were collected from different locations in the study area and analyzed for fluoride levels using a spectrophotometer. The fluoride levels range from 0.6 to 1.8 mg/L, with an average value of 1.31 mg/L. Overall, 32% of groundwater samples found to be exceeded the prescribed limits of fluoride in the study area, as prescribed by the World Health Organization (WHO). Besides, the optimal amounts of fluoride in drinking water have to meet in the range of 0.5 mg/L and 1.5 mg/L, as this range is the permissible limit for drinking. Fluoride is an essential micronutrient for normal mineralization of teeth and bones as well as for the formation of dental enamel in the human body. This study suggested that prolonged intake of drinking water with fluoride concentrations higher than acceptable limits can trigger dental and skeletal fluorosis in the general community of the study region. The fluoride data compared with the permissible levels recommended for drinking purposes by various regulatory agencies across the world.

Chryseomicrobium palamuruense sp. nov., a haloalkalitolerant bacterium isolated from a sediment sample.

A novel Gram-stain-positive, rod shaped, motile bacterium, designated strain PU1T, was isolated from a sediment sample collected from a drainage near hostel of Palamuru University, Mahabubnagar district, T.S, India (16°43'23″N 77°58'49″E). Cells of strain PU1T are positive for catalase, oxidase, phosphatase, lipase and urease, and negative for gelatinase, amylase, protease, cellulase, lysine decarboxylase and ornithine decarboxylase. The fatty acids were dominated by saturated fatty acids (82.7 %), with a high abundance of iso-C15 : 0 (48.8 %), anteiso-C15 : 0 (7.3 %), iso-C16 : 0 (11.9 %), C16 : 1ω7c alcohol (11.8 %) and iso-C17 : 0 (5.3 %). Strain PU1T contained MK-8 as the major respiratory quinone and diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine make up the phospholipid composition. The cell-wall peptidoglycan contains meso-diaminopimelic acid as the diamino acid and cell-wall sugars are d-glucose and d-galactose. 16S rRNA gene sequence analysis indicated Chryseomicrobium imtechense and Chryseomicrobiumamylolyticum, members of family Planococcaceae within the phylum Firmicutes, are the closest related species with 16S rRNA gene sequence similarities of 99 %. Other members of the family Planococcaceae had sequence similarities of 99 %, and DNA-DNA relatedness values between strain PU1T and Chryseomicrobium imtechense MW 10T, Chryseomicrobiumamylolyticum JC16T were 38 and 32 % respectively. The G+C content of DNA of strain PU1T is 48.5 mol%. Based on the above-mentioned phenotypic and phylogenetic characteristics, strain PU1T represents a novel species of the genus Chryseomicrobium for which the name Chryseomicrobiumpalamuruense sp. nov. is proposed. The type strain is PU1T(=CCUG 59101T=JCM 16712T=KCTC 13722T=NBRC106750T).

Isolation and characterization of novel lipase gene LipHim1 from the DNA isolated from soil samples.

Metagenomics is a magnificent tool to isolate genes from unknown/uncharacterized species and also from organisms that cannot be cultured. In this study, we constructed a metagenomic library from isolated DNA of soil samples collected from Palamuru University campus premises, in Mahabubnagar district of Andhra Pradesh, India. We isolated a novel lipase gene LipHim1, which has an open reading frame of 591 base pairs and encodes ∼23 kDa protein consisting of 196 amino acids. The Lipase LipHim1 showed maximum 32% homology at the protein level with the extracellular Aeromonas hydrophila lipase (Class II, GDSL family) and was significantly different from all other known lipases. The isolated lipase catalyzed the hydrolysis of fatty acid esters of polyoxyethylene sorbitan such as Tween 60. Our results indicate that the isolated lipase gene is novel.

Plant growth regulation of Bt-cotton through Bacillus species.

Deccan plateau in India periodically experiences droughts due to irregular rain fall and the soil in many parts of the region is considered to be poor for farming. Plant growth promoting rhizobacteria are originally defined as root-colonizing bacteria, i.e., Bacillus that cause either plant growth promotion or biological control of plant diseases. The study aims at the isolation of novel Bacillus species and to assess the biotechnological potential of the novel species as a biofertilizer, with respect to their plant growth promoting properties as efficient phosphate-solubilizing bacteria. Seven different strains of Bacillus were isolated from cotton rhizosphere soil near boys' hostel of Palamuru University which belongs to Deccan plateau. Among seven isolated strains, Bacillus strain-7 has shown maximum support for good growth of eight cotton cultivars. This bacterial species is named Bacillus sp. PU-7 based on the phenotypic and phylogenetic analysis. Among eight cotton cultivars, Mahyco has shown high levels of IAA, proteins, chlorophyll, sugars and low level of proline. Efficacy of novel Bacillus sp. PU-7 with Mahyco cultivar has been checked experimentally at field level in four different cotton grown agricultural soils. The strains supported plant growth in almost all the cases, especially in the deep black soil, with a clear evidence of maximum plant growth by increased levels of phytohormone production and biochemical analysis, followed by shallow black soil. Hence, it is inferred that the novel isolate can be used as bioinoculant in the cotton fields.

Identification of opportunistic pathogenic bacteria in drinking water samples of different rural health centers and their clinical impacts on humans.

International drinking water quality monitoring programs have been established in order to prevent or to reduce the risk of contracting water-related infections. A survey was performed on groundwater-derived drinking water from 13 different hospitals in the Mahabubnagar District. A total of 55 bacterial strains were isolated which belonged to both gram-positive and gram-negative bacteria. All the taxa were identified based on the 16S rRNA gene sequence analysis based on which they are phylogenetically close to 27 different taxa. Many of the strains are closely related to their phylogenetic neighbors and exhibit from 98.4 to 100% sequence similarity at the 16S rRNA gene sequence level. The most common group was similar to Acinetobacter junii (21.8%) and Acinetobacter calcoaceticus (10.9%) which were shared by 7 and 5 water samples, respectively. Out of 55 isolates, only 3 isolates belonged to coliform group which are Citrobacter freundii and Pantoea anthophila. More than half (52.7%, 29 strains) of the phylogenetic neighbors which belonged to 12 groups were reported to be pathogenic and isolated from clinical specimens. Out of 27 representative taxa are affiliated have eight representative genera in drinking water except for those affiliated with the genera Exiguobacterium, Delftia, Kocuria, and Lysinibacillus.

Vertical distribution of bacteria in a lake sediment from Antarctica by culture-independent and culture-dependent approaches.

Bacterial diversity of the subsurface (18-22 cm), middle (60-64 cm) and bottom (100-104 cm) of a 136-cm-long sediment core sampled from a freshwater lake in Antarctica was determined by the culturable approach, T-RFLP and 16S rRNA gene clone libraries. Using the culturable approach, 41 strains were isolated and, based on phylogenetic analysis, they could be categorized into 14 groups. Representatives of the 14 groups varied in their growth temperature range (4-30 °C), in their tolerance to NaCl (0-2 M NaCl) and in the growth pH range (5-11). Eleven of fourteen representative strains exhibited either amylase, lipase, protease and (or) urease activities at 4 °C. Bacterial diversity at the phyla level using T-RFLP and 16S rRNA clone libraries was similar and clones were affiliated with Proteobacteria, Bacteroidetes, Actinobacteria and Firmicutes. TRFs affiliated with Spirochaetes were detected only by the T-RFLP approach and clones affiliated with Caldiserica only in the clone libraries. Stratification of bacteria along the depth of the sediment was observed both with the T-RFLP and the 16S rRNA gene clone library methods, and results indicated that stratification was dependent on the nature of the organism, aerobic or anaerobic. For instance, aerobic Janthinobacterium and Polaromonas were confined to the surface of the sediment, whereas anaerobic Caldisericum was present only in the bottom portion of the core. It may be concluded that the bacterial diversity of an Antarctic lake sediment core sample varies throughout the length of the core depending on the oxic-anoxic conditions of the sediment. Furthermore, these psychrophilic bacteria, due to their ability to produce extracellular cold active enzymes, might play a key role in the transformation of complex organic compounds.

Bacterial biodiversity from Roopkund Glacier, Himalayan mountain ranges, India.

The bacterial diversity of two soil samples collected from the periphery of the Roopkund glacial lake and one soil sample from the surface of the Roopkund Glacier in the Himalayan ranges was determined by constructing three 16S rRNA gene clone libraries. The three clone libraries yielded a total of 798 clones belonging to 25 classes. Actinobacteria was the most predominant class (>10% of the clones) in the three libraries. In the library from the glacial soil, class Betaproteobacteria (24.2%) was the most predominant. The rarefaction analysis indicated coverage of 43.4 and 41.2% in the samples collected from the periphery of the lake thus indicating a limited bacterial diversity covered; at the same time, the coverage of 98.4% in the glacier sample indicated most of the diversity was covered. Further, the bacterial diversity in the Roopkund glacier soil was low, but was comparable with the bacterial diversity of a few other glaciers. The results of principal component analysis based on the 16S rRNA gene clone library data, percentages of OTUs and biogeochemical data revealed that the lake soil samples were different from the glacier soil sample and the biogeochemical properties affected the diversity of microbial communities in the soil samples.

Arthrobacter antarcticus sp. nov., isolated from an Antarctic marine sediment.

A bacterial strain, SPC26(T), was isolated from a sediment sample of the Southern Ocean off Antarctica. The strain was Gram-staining- and catalase-positive and contained lysine and alanine in the cell-wall peptidoglycan. The major cellular fatty acids were anteiso-C15:0 (54.92 %), iso-C15:0 (11.47 %), anteiso-C17:0 (6.48 %) and anteiso-C15:1 (6.38 %) and the major menaquinones were MK-8, MK-9 and MK-10. The major polar lipids were phosphatidylethanolamine and diphosphatidylglycerol. The G+C content was 68 ± 0.5 mol%. Based on 16S rRNA gene sequence similarities, the nearest phylogenetic neighbours of strain SPC26(T) were identified as Arthrobacter gangotriensis Lz1y(T) (98.8 %), A. sulfureus DSM 20167(T) (98.6 %), A. psychrophenolicus DSM 15454(T) (97.9 %) and A. kerguelensis KGN15(T) (97.5). With these strains, strain SPC26(T) exhibited DNA-DNA relatedness values of 36, 21, 12 and 10 %, respectively. Therefore, on the basis of 16S rRNA gene sequence comparisons, phylogenetic analysis, phenotypic characteristics and DNA-DNA relatedness, it is proposed that strain SPC26(T) represents a novel species of Arthrobacter, for which the name Arthrobacter antarcticus sp. nov. is proposed, with strain SPC26(T) (=LMG 24542(T) =NCCB 100228(T)) as the type strain.

Janibacter hoylei sp. nov., Bacillus isronensis sp. nov. and Bacillus aryabhattai sp. nov., isolated from cryotubes used for collecting air from the upper atmosphere.

Three novel bacterial strains, PVAS-1(T), B3W22(T) and B8W22(T), were isolated from cryotubes used to collect air samples at altitudes of between 27 and 41 km. Based on phenotypic characteristics, chemotaxonomic features, DNA-DNA hybridization with the nearest phylogenetic neighbours and phylogenetic analysis based on partial 16S rRNA gene sequences (PVAS-1(T), 1196 nt; B3W22(T), 1541 nt; B8W22(T), 1533 nt), the three strains were identified as representing novel species, and the names proposed are Janibacter hoylei sp. nov. (type strain PVAS-1(T) =MTCC 8307(T) =DSM 21601(T) =CCUG 56714(T)), Bacillus isronensis sp. nov. (type strain B3W22(T) =MTCC 7902(T) =JCM 13838(T)) and Bacillus aryabhattai sp. nov. (type strain B8W22(T) =MTCC 7755(T) =JCM 13839(T)).

Description of Leifsonia kafniensis sp. nov. and Leifsonia antarctica sp. nov.

Strains KFC-22(T) and SPC-20(T) are yellow-pigmented, Gram-positive, aerobic, non-motile, rod-shaped bacteria that were isolated from a soil sample near the Kafni glacier in the Himalayan mountain ranges in India, and from a spade core sediment sample from the Antarctic Ocean at Larsemann Hill, respectively. In both cases, the cell-wall peptidoglycan contained 2,4-diaminobutyric acid as the diamino acid, anteiso-C(15 : 0), anteiso-C(17 : 0) and iso-C(16 : 0) were the predominant fatty acids and MK-11 was the major isoprenoid quinone in the cell membrane. On the basis of the above-mentioned characteristics, both strains can be assigned to the genus Leifsonia. The strains share 16S rRNA gene sequence similarity of 97.7 % and DNA relatedness of only 10 %, indicating that they represent different species. A blast analysis indicated that Leifsonia pindariensis PON10(T) was the closest phylogenetic neighbour of strains SPC-20(T) and KFC-22(T), showing 16S rRNA gene sequence similarities of 97.3 and 97.7 %, respectively. However, at the whole-genome level, strains KFC-22(T) and SPC-20(T) shared 42 and 11 % DNA-DNA relatedness, respectively, with L. pindariensis PON10(T). In addition, both strains exhibited several phenotypic differences with respect to L. pindariensis PON10(T). Thus, on the basis of the differences that the two strains exhibited with respect to L. pindariensis, both were identified as representing novel species of the genus Leifsonia, for which the names Leifsonia kafniensis sp. nov. (type strain KFC-22(T) =NCCB 100216(T) =LMG 24362(T)) and Leifsonia antarctica sp. nov. (type strain SPC-20(T) =NCCB 100227(T) =LMG 24541(T)) are proposed.

Importance of trmE for growth of the psychrophile Pseudomonas syringae at low temperatures.

Transposon mutagenesis of Pseudomonas syringae Lz4W, a psychrophilic bacterium capable of growing at temperatures between 2 and 30 degrees C, yielded 30 cold-sensitive mutants, and CSM1, one of these cold-sensitive mutants, was characterized. Growth of CSM1 was retarded when it was cultured at 4 degrees C but not when it was cultured at 22 degrees C and 28 degrees C compared to the growth of wild-type cells, indicating that CSM1 is a cold-sensitive mutant of P. syringae Lz4W. The mutated gene in CSM1 was identified as trmE (coding for tRNA modification GTPase), and evidence is provided that this gene is induced at low temperatures. Further, the cold-inducible nature of the trmE promoter was demonstrated. In addition, the transcription start site and the various regulatory elements of the trmE promoter, such as the -10 region, -35 region, UP element, cold box, and DEAD box, were identified, and the importance of these regulatory elements in promoter activity were confirmed. The importance of trmE in rapid adaptation to growth at low temperatures was further highlighted by plasmid-mediated complementation that alleviated the cold-sensitive phenotype of CSM1.

Exiguobacterium soli sp. nov., a psychrophilic bacterium from the McMurdo Dry Valleys, Antarctica.

A novel psychrophilic bacterium, designated strain DVS 3Y(T), was isolated from a moraine sample from the McMurdo Dry Valleys, Antarctica. Phenotypic and chemotaxonomic characteristics and data from a phylogenetic analysis based on 16S rRNA gene sequences indicated that strain DVS 3Y(T) was related to the genus Exiguobacterium. Strain DVS 3Y(T) exhibited >97 % gene sequence similarity with respect to Exiguobacterium acetylicum DSM 20416(T) (97.4 %), Exiguobacterium oxidotolerans DSM 17272(T) (97.4 %), Exiguobacterium indicum IAM 15368(T) (97.6 %), Exiguobacterium undae DSM 14481(T) (98.2 %), Exiguobacterium sibiricum DSM 17290(T) (98.6 %) and Exiguobacterium antarcticum DSM 14480(T) (99.6 %). However, the levels of DNA-DNA relatedness between strain DVS 3Y(T) and E. acetylicum DSM 20416(T), E. oxidotolerans JCM 12280(T), E. indicum IAM 15368(T), E. undae DSM 14481(T), E. sibiricum DSM 17290(T) and E. antarcticum DSM 14480(T) were less than 70 % (52, 25, 16, 33, 16 and 38 %, respectively). Strain DVS 3Y(T) also differed from these six closely related species in a number of phenotypic traits. Phenotypic, chemotaxonomic and phylogenetic data suggest that strain DVS 3Y(T) represents a novel species of the genus Exiguobacterium, for which the name Exiguobacterium soli sp. nov. is proposed. The type strain is DVS 3Y(T) (=MTCC 4816(T)=JCM 14376(T)).

Brevibacterium oceani sp. nov., isolated from deep-sea sediment of the Chagos Trench, Indian Ocean.

Two bacterial strains, designated BBH5 and BBH7(T), were isolated from a deep-sea sediment sample collected from the Chagos Trench of the Indian Ocean (1 degrees 06' S 7 degrees 31' E). Based on their 16S rRNA gene sequence similarity (99.9%), level of DNA-DNA relatedness (93%) and a number of similar phenotypic characteristics, the two strains are identified as representing the same species. Their phylogenetically nearest neighbours, based on 16S rRNA gene sequence similarity values (97.9-98.4%), were identified as Brevibacterium iodinum, Brevibacterium epidermidis, Brevibacterium linens and Brevibacterium permense. However, strains BBH5 and BBH7(T) could be distinguished from the above four species by a number of phenotypic characteristics, and levels of DNA-DNA relatedness between the two new isolates and these Brevibacterium species were 35-42%. Therefore, strains BBH5 and BBH7(T) are considered to represent a novel species of the genus Brevibacterium, for which the name Brevibacterium oceani sp. nov. is proposed. The type strain is BBH7(T) (=LMG 23457(T) =IAM 15353(T)).

Microbacterium indicum sp. nov., isolated from a deep-sea sediment sample from the Chagos Trench, Indian Ocean.

Two bacterial strains, BBH6(T) and BBH9, were isolated from a deep-sea sediment sample collected from the Chagos Trench, Indian Ocean, at a depth of 5904 m. The two strains were closely related in their 16S rRNA gene sequences (99.7 %), belonged to one genomic species and were virtually identical at the phenotypic level. Microbacterium barkeri DSM 20145(T) was the nearest phylogenetic neighbour to the new isolates, with 16S rRNA gene sequence similarity levels of 97.2-97.4 %. The new isolates exhibited levels of DNA-DNA relatedness of 32-34 % to M. barkeri and differed from it in a number of phenotypic characteristics. Therefore, it is suggested that strains BBH6(T) and BBH9 represent a novel species of the genus Microbacterium, for which the name Microbacterium indicum sp. nov. is proposed. The type strain is BBH6(T) (=LMG 23459(T)=IAM 15355(T)).