Structural investigations into the binding mode of novel neolignans Cmp10 and Cmp19 microtubule stabilizers by in silico molecular docking,molecular dynamics,and binding free energy calculations

Microtubule stabilizers provide an important mode of treatment via mitotic cell arrest of cancer cells. Recently, we reported two novel neolignans derivatives Cmp10 and Cmp19 showing anticancer activity and working as microtubule stabilizers at micromolar concentrations. In this study, we have explored the binding site, mode of binding, and stabilization by two novel microtubule stabilizers Cmp10 and Cmp19 using in silico molecular docking, molecular dynamics (MD) simulation, and binding free energy calculations. Molecular docking studies were performed to explore the β-tubulin binding site of Cmp10 and Cmp19. Further, MD simulations were used to probe the β-tubulin stabilization mechanism by Cmp10 and Cmp19. Binding affinity was also compared for Cmp10 and Cmp19 using binding free energy calculations. Our docking results revealed that both the compounds bind at Ptxl binding site in β-tubulin. MD simulation studies showed that Cmp10 and Cmp19 binding stabilizes M-loop (Phe272-Val288) residues of β-tubulin and prevent its dynamics, leading to a better packing between α and β subunits from adjacent tubulin dimers. In addition, His229, Ser280 and Gln281, and Arg278, Thr276, and Ser232 were found to be the key amino acid residues forming H-bonds with Cmp10 and Cmp19, respectively. Consequently, binding free energy calculations indicated that Cmp10 (?113.655 kJ/mol) had better binding compared to Cmp19 (?95.216 kJ/mol). This study provides useful insight for better understanding of the binding mechanism of Cmp10 and Cmp19 and will be helpful in designing novel microtubule stabilizers.     

MMP-9 gene ablation mitigates hyperhomocystenemia-induced cognition and hearing dysfunction

Hyperhomocysteinemia (HHcy) is associated with cognitive decline and hearing loss due to vascular dysfunction. Although we have shown that HHcy-induced increased expression of matrix metalloproteinase-9 (MMP-9) is associated with cochlear pathology in cystathionine-β-synthase heterozygous (CBS^+/?) mice, it is still unclear whether MMP-9 contributes to functional deficit in cognition and hearing. Therefore, we hypothesize that HHcy-induced MMP-9 activation causes vascular, cerebral and cochlear remodeling resulting in diminished cognition and hearing. Wildtype (WT), CBS^+/?, MMP-9^?/? and CBS^+/?/MMP-9^?/? double knock-out (DKO) mice were genotyped and used. Doppler flowmetry of internal carotid artery (ICA) was performed for peak systolic velocity [PSV], pulsatility index [PI] and resistive index [RI]. Cognitive functions were assessed by Novel Object Recognition Test (NORT) and for cochlear function Auditory brainstem response (ABR) was elicited. Peak systolic velocity, pulsatility and resistive indices of ICA were decreased in CBS^+/? mice, indicating reduced perfusion. ABR threshold was increased and maximum ABR amplitude and NORT indices (recognition, discrimination) were decreased in CBS^+/? mice compared to WT and MMP-9^?/?. All these parameters were attenuated in DKO mice suggesting a significant role of MMP-9 in HHcy-induced vascular, neural and cochlear pathophysiology. Regression analysis of PSV with ABR and cognitive parameters revealed significant correlation (0.44–0.58). For the first time, MMP-9 has been correlated directly to functional deficits of brain and cochlea, and found to have a significant role. Our data suggests a dual pathology of HHcy occurring due to a decrease in blood supply (vasculo-neural and vasculo-cochlear) and direct tissue remodeling.     

A new family of dispersed repeats from Brassica nigra: characterization and localization

The 459-bp HindIII (pBN-4) and the 1732-bp Eco RI (pBNE8) fragments from the Brassica nigra genome were cloned and shown to be members of a dispersed repeat family. Of the three major diploid Brassica species, the repeat pBN-4 was found to be highly specific for the B. nigra genome. The family also hybridized to Sinapis arvensis showing that B. nigra had a closer relationship with the S. arvensis genome than with B. oleracea or B. campestris. The clone pBNE8 showed homology to a number of tRNA species indicating that this family of repeats may have originated from a tRNA sequence. The species-specific 459-bp repeat pBN-4 was localized on the B. nigra chromosomes using monosomic addition lines. In addition to the localization of pBN-4, the chromosomal distribution of two other species-specific repeats, pBN34 and pBNBH35 (reported earlier), was studied. The dispersed repeats pBN-4 and pBNBH35 were found to be present on all of the chromosomes, whereas the tandem repeat pBN34 was localized on two chromosomes.     

Plant responses to water stress: changes in growth,dry matter production,stomatal frequency and leaf anatomy

The responses of seedlings of three fast growing tree species,Eucalyptus hybrid(E. camaldulensis × E. teriticornis), Casuarina equisetifolia andMelia azedarach, to different levels of soil moisture in controlled glasshouse conditions were compared. The survival percentage, height of plants, number of leaves per plant, number of branches, fresh mass and dry mass of roots, stems, branches and leaves decreased in the three species with increasing water stress. Stomatal frequency and length of stomata inEucalyptus andMelia also decreased with increasing water stress. However, no significant difference was obtained in the width of stomata and the ratio of number of open stomata to total number of stomata per unit area. The leaf thickness decreased, but the thickness of palisade parenchyma increased with increasing water stress inEucalyptus hybrid andCasuarina. Leaf thickness ofMelia did not show any significant variation due to water stress.     

Role of Liv-52 in protection against beryllium intoxication

An ayurvedic medicine, Liv-52, was studied as a prophylactic agent against beryllium-induced toxicity in rats. Administration of berylliumper se caused severe degenerative and necrotic changes in kidneys, liver, and uterus. Beryllium exposure also reduced glycogen content, activities of alkaline phosphatase, succinate-dehydrogenase, and adenosine-triphosphatase in these organs. On the contrary, activities of acid phosphatase and glucose-6-phosphatase showed marginal increase. Liv-52-primed rats exhibited comparatively less marked toxic effects.     

Antioxidant and Antidiabetic Properties of Extracts from Three Underutilized Food Plants of North East India

Three underutilized leafy vegetables Sarcochlamys pulcherrima (Roxb.) Gaudich (SP), Ipomoea aquatica Forssk. (IA) and Zanthoxylum rhetsa (Roxb.) DC (ZR) were extracted with different solvents viz. 95 % ethyl alcohol, methanol and hot water. The extracts were evaluated for their antioxidant potential via DPPH, ABTS and FRAP assay along with electroanalytical studies using cyclic voltammetry. The antidiabetic potential was determined by recording their α-amylase and α-glucosidase inhibitory assay. The total phenolic content (TPC), total flavonoid content (TFC) and the liquid chromatography-mass spectrometry (LC/MS) based phytochemical profiles of the extracts were also determined. All three extracts of SP exhibited significant antioxidant capacity. The antidiabetic potential of the IA and ZR extracts was found to be higher than or at par with that of standard acarbose. LC/MS studies reveal the presence of hitherto reported antioxidant and antidiabetic compounds like gamma-aminobutyric acid, cinnamic acid, caffeic acid, α-viniferin, piperlonguminine, niacin, kaempferol, etc., in the extracts.     

Shoot growth phenology of co-existing evergreen and deciduous species in an oak forest

Shoot growth phenology was compared for the saplings of evergreen and deciduous woody species sharing the same microsite. Growth initiation occurred earlier in evergreens (among co-stratal species) while deciduous species completed their growth earlier. Shoot growth rate was significantly greater (P<0.01) for deciduous trees than evergreen trees. The amount of shoot elongations and shoot diameter was also significantly greater (P<0.01) for deciduous trees than evergreens. On the other hand, among shrubs the amount of shoot elongation and shoot diameter was greater for evergreens but the rate of elongation and diameter was more or less similar for both. The duration of shoot elongation and shoot diameter was significantly longer in evergreens than the deciduous species. Leaf packing (number of leaves per shoot) was significantly more dense in evergreen trees (P<0.01) than in deciduous tree species. Leaf packing was more dense in evergreen than deciduous shrubs but the difference was not significant. Leaf area (per individual leaf) at full expansion was significantly greater (P<0.01) in deciduous species. Leaf dry mass and specific leaf mass in the initial stage was significantly greater for evergreen species than for deciduous species. The number of buds/10 cm of shoot was higher in evergreens. However, the per cent mortality was also higher in them.     

Keratinolytic proteinase produced by Candida albicans

Candida albicans was cultivated in various media that contained human stratum corneum, human scalp hair or keratin powder (cow's hoof) as a nitrogen source. Production of a keratinolytic proteinase (KPase) was observed when C. albicans was incubated in the medium containing stratum corneum. However, there was no production of a KPase that could digest human stratum corneum in the medium containing hair or keratin powder. alpha-fibrous protein extracted from human stratum corneum was digested by the KPase. The pH optimum of the enzyme was 4.0 and enzyme activity was inhibited by pepstatin A and chymostatin. The KPase, a kind of carboxyl proteinase, may be important for C. albicans to enable it to play a pathogenic role in vivo.     

Studies on a strain of Fusarium solani (Mart.) Sacc. Isolated from a case of mycotic keratitis

A strain of Fusarium solani (Mart.) Sacc. (IMI-216517), isolated from a patient of mycotic keratitis, produced experimental keratomycosis in albino rabbit cornea and survived in internal tissues of albino mice for varying periods. Alantolactone, isolated from the plant — Inula racemosa Hook. f. exhibited marked in vitro fungistatic activity against this strain of F. solani at 100–200 g/ml concentrations. The strain was less sensitive to amphotericin B and showed more acid than alkaline proteinase and phosphatase activities.Communication No. 2526.