Production of lipase of Aspergillus foetidus in a batch stirred reactor

Summary Maximum lipase production byAspergillus foetidus was obtained from cultures grown in the medium of 2% olive oil and 0.5% sucrose. The optimal conditions for the production of lipases in the Multigen fermenters were found to be at 500rpm with an airflow of 1.5 liter per mimute. Immobilization of the fungal source was found to be infeasible in natural polymers.     

A histone 1-like antigen is a component of the nuclear envelope

Rabbit antibodies have been raised against rat liver nuclear envelopes. An enzyme-linked immunosorbent assay (ELISA) demonstrated high titer antiserum specific for the nuclear envelope preparation. Immunocytochemical studies showed that the antiserum stained the nuclear envelopes, but not intra-nuclear components of HEp-2 (human malignant epithelial) cells. When electrophoretically separated peptides were tested by immunoblotting techniques, the rabbit antiserum specifically stained proteins with molecular masses of 26 and 28 kD. These peptides had similar mobilities to purified histone 1 (H1). Indeed purified calf thymus H1 recognized the antiserum. The antigens are not loosely bound to the nuclear envelope, as they could not be extracted with low salt. Therefore, we have established that the 26 and 28 kD nuclear envelope peptides are not contaminants of the nuclear envelope preparation and that they express determinants that are immunologically cross-reactive with purified H1, but not with intra-nuclear H1.     

Diet fat alters the structure and function of the nuclear envelope: modulation of membrane fatty acid composition, NTPase activity and binding of triiodothyronine

Mice were fed a diet either high or low in P/S ratio to determine the effect of altering dietary lipid on the fatty acid composition of liver nuclear envelopes and thereby on functions of the nuclear envelope. Mice fed the high P/S diet exhibited higher levels of C18:2 omega 6 and unsaturates in liver nuclear envelopes, higher specific activity of NTPase and specific binding for L-triiodothyronine at 15 degrees C and 22 degrees C compared with the low P/S diet fed group. These observations indicate that diet-induced differences in the fatty acid composition of nuclear envelope lipid affects functions of the nuclear envelope.     

Insulin binding to liver nuclei from lean and obese mice is altered by dietary fat.

Insulin binding to the plasma membrane is known to be altered by modifying the membrane composition through dietary treatment. As insulin binding receptors are also present on nuclear membrane, this study was undertaken to investigate if specific binding of insulin to the liver nuclei is altered by diet. 8-wk-old female C57 B 6J lean and ob/ob mice were fed semipurified diets containing 20% (w/w) fat of either high or low polyunsaturated-to-saturated (P/S) fatty acid ratio for 4 wk. Liver nuclei were prepared, insulin binding was measured and nuclear phospholipids were isolated for lipid analysis. Insulin binding was highest in nuclei prepared from lean mice fed a high P/S diet. Specific binding of insulin to nuclei prepared from obese mice was also increased by the high P/S diet, but to a lesser extent compared to lean mice. Feeding a high P/S diet increased polyunsaturated fatty acid content of membrane phospholipids from both lean and ob/ob mice. Obese mice were characterized by higher levels of arachidonic acid and lower levels of linoleic acid in phosphatidylcholine. The present study establishes that insulin binding to liver nuclei is increased by feeding a high P/S diet, and that insulin binding to liver nuclei from obese mice is lower than from lean mice.     

Production of ergot alkaloids by Aspergillus fumigatus (Fresenius)

Summary Nutritional requirements for the production of ergot alkaloids were studied with Aspergillus fumigatus under submerged conditions of fermentation, in a chemically defined medium. Glucose in combination with mannitol and triammonium citrate were found to be the best sources of carbon and nitrogen for the production of alkaloids. Carbon to nitrogen ratio of 4.16 : 1 was found optimum. Phosphate at elevated concentration inhibited alkaloid production.     

A Biodegradable,Sustained-Released,Prednisolone Acetate Microfilm Drug Delivery System Effectively Prolongs Corneal Allograft Survival in the Rat Keratoplasty Model

Frequent and long-term use of topical corticosteroids after corneal transplantation is necessary to prevent graft rejection. However, it relies heavily on patient compliance, and sustained therapeutic drug levels are often not achieved with administration of topical eye drops. A biodegradable drug delivery system with a controlled and sustained drug release may circumvent these limitations. In this study, we investigated the efficacy of a prednisolone acetate (PA)-loaded poly (d,l-lactide-co-ε-caprolactone) (PLC) microfilm drug delivery system on promoting the survival of allogeneic grafts after penetrating keratoplasty (PK) using a rat model. The drug release profiles of the microfilms were characterized (group 1). Subsequently, forty-eight PK were performed in four experimental groups: syngeneic control grafts (group 2), allogeneic control grafts (group 3), allogeneic grafts with subconjunctivally-implanted PA microfilm (group 4), and allogeneic grafts with PA eye drops (group 5; n = 12 in each). PA-loaded microfilm achieved a sustained and steady release at a rate of 0.006–0.009 mg/day, with a consistent aqueous drug concentration of 207–209 ng/ml. The mean survival days was >28 days in group 2, 9.9±0.8 days in group 3, 26.8±2.7 days in group 4, and 26.4±3.4 days in group 5 (P = 0.023 and P = 0.027 compared with group 3). Statistically significant decrease in CD4+, CD163+, CD 25+, and CD54+ cell infiltration was observed in group 4 and group 5 compared with group 3 (P<0.001). There was no significant difference in the mean survival and immunohistochemical analysis between group 4 and group 5. These results showed that sustained PA-loaded microfilm effectively prolongs corneal allograft survival. It is as effective as conventional PA eye drops, providing a promising clinically applicable alternative for patients undergoing corneal transplantation.     

Origin and diversification of Indian Ceropegieae (Apocynaceae) and its possible relation to the Indian monsoon

The Indian subcontinent has experienced a major shift in climatic regime from a wet tropical regime to increased seasonal rainfall, since the late Miocene. This shift has been attributed to the intensification of monsoons, which led to opening up of dry habitats in humid forests and formation of deciduous forests. We explored the role of this climatic shift in the origin and diversification of dry‐adapted plant genera Ceropegia and Brachystelma (Ceropegiae, Asclepiadoideae, Apocynaceae). We sampled Ceropegia and Brachystelma from across India and used five markers (two nuclear and three plastid regions) to reconstruct a global phylogeny of this group. Indian members of the tribe Ceropegiae were derived from Africa through at least four independent dispersal events. All dispersal events occurred in late Miocene after establishment of a monsoon climate. One of these early dispersing lineages underwent rapid radiation in peninsular India, giving rise to around 50 species. Thus, both dispersal and diversification events coincided with the intensification of monsoons and concomitant aridification. The role of environment in the evolution of floral characteristics and root type in the Indian radiation is also discussed. This is one of the first reports on a dry‐adapted endemic radiation of plants in India.     

Design,Synthesis, Evaluation and Thermodynamics of 1-Substituted Pyridylimidazo[1,5-a]Pyridine Derivatives as Cysteine Protease Inhibitors

Targeting papain family cysteine proteases is one of the novel strategies in the development of chemotherapy for a number of diseases. Novel cysteine protease inhibitors derived from 1-pyridylimidazo[1,5-a]pyridine representing pharmacologically important class of compounds are being reported here for the first time. The derivatives were initially designed and screened in silico by molecular docking studies against papain to explore the possible mode of action. The molecular interaction between the compounds and cysteine protease (papain) was found to be very similar to the interactions observed with the respective epoxide inhibitor (E-64c) of papain. Subsequently, compounds were synthesized to validate their efficacy in wet lab experiments. When characterized kinetically, these compounds show their K_i and IC₅₀ values in the range of 13.75 to 99.30 µM and 13.40 to 96.50 µM, respectively. The thermodynamics studies suggest their binding with papain hydrophobically and entropically driven. These inhibitors also inhibit the growth of clinically important different types of Gram positive and Gram negative bacteria having MIC₅₀ values in the range of 0.6–1.4 µg/ml. Based on Lipinski’s rule of Five, we also propose these compounds as potent antibacterial prodrugs. The most active antibacterial compound was found to be 1-(2-pyridyl)-3-(2-hydroxyphenyl)imidazo[1,5-a]pyridine (3a).     

The Human Eye Proteome Project: Perspectives on an emerging proteome

There are an estimated 285 million people with visual impairment worldwide, of whom 39 million are blind. The pathogenesis of many eye diseases remains poorly understood. The human eye is currently an emerging proteome that may provide key insight into the biological pathways of disease. We review proteomic investigations of the human eye and present a catalogue of 4842 nonredundant proteins identified in human eye tissues and biofluids to date. We highlight the need to identify new biomarkers for eye diseases using proteomics. Recent advances in proteomics do now allow the identification of hundreds to thousands of proteins in tissues and fluids, characterization of various PTMs and simultaneous quantification of multiple proteins. To facilitate proteomic studies of the eye, the Human Eye Proteome Project (HEPP) was organized in September 2012. The HEPP is one of the most recent components of the Biology/Disease‐driven Human Proteome Project (B/D‐HPP) whose overarching goal is to support the broad application of state‐of‐the‐art measurements of proteins and proteomes by life scientists studying the molecular mechanisms of biological processes and human disease. The large repertoire of investigative proteomic tools has great potential to transform vision science and enhance understanding of physiology and disease processes that affect sight.     

Identification of Pak1 inhibitors using water thermodynamic analysis

Abstract

p21-activated kinases (Paks) play an integral component in various cellular diverse processes. The full activation of Pak is dependent upon several serine residues present in the N-terminal region, a threonine present at the activation loop, and finally the phosphorylation of these residues ensure the complete activation of Pak1. The present study deals with the identification of novel potent candidates of Pak1 using computational methods as anti-cancer compounds. A diverse energy based pharmacophore (e-pharmacophore) was developed using four co-crystal inhibitors of Pak1 having pharmacophore features of 5 (DRDRR), 6 (DRHADR), and 7 (RRARDRP and DRRDADH) hypotheses. These models were used for rigorous screening against e-molecule database. The obtained hits were filtered using ADME/T and molecular docking to identify the high affinity binders. These hits were subjected to hierarchical clustering using dendritic fingerprint inorder to identify structurally diverse molecules. The diverse hits were scored against generated water maps to obtain WM/MM ΔG binding energy. Furthermore, molecular dynamics simulation and density functional theory calculations were performed on the final hits to understand the stability of the complexes. Five structurally diverse novel Pak1 inhibitors (4835785, 32198676, 32407813, 76038049, and 32945545) were obtained from virtual screening, water thermodynamics and WM/MM ΔG binding energy. All hits revealed similar mode of binding pattern with the hinge region residues replacing the unstable water molecules in the binding site. The obtained novel hits could be used as a platform to design potent drugs that could be experimentally tested against cancer patients having increased Pak1 expression.