Parameter determination and validation for a mechanistic model of the enzymatic saccharification of cellulose‐Iβ

Cost‐effective production of fuels and chemicals from lignocellulosic biomass often involves enzymatic saccharification, which has been the subject of intense research and development. Recently, a mechanistic model for the enzymatic saccharification of cellulose has been developed that accounts for distribution of cellulose chain lengths, the accessibility of insoluble cellulose to enzymes, and the distinct modes of action of the component cellulases [Griggs et al. (2012) Biotechnol. Bioeng., 109(3):665–675; Griggs et al. (2012) Biotechnol. Bioeng., 109(3):676–685]. However, determining appropriate values for the adsorption, inhibition, and rate parameters required further experimental investigation. In this work, we performed several sets of experiments to aid in parameter estimation and to quantitatively validate the model. Cellulosic materials differing in degrees of polymerization and crystallinity (α‐cellulose‐I_β and highly crystalline cellulose‐I_β) were digested by component enzymes (EG_I/CBH_I/ ) and by mixtures of these enzymes. Based on information from the literature and the results from these experiments, a single set of model parameters was determined, and the model simulation results using this set of parameters were compared with the experimental data of total glucan conversion, chain‐length distribution, and crystallinity. Model simulations show significant agreement with the experimentally derived glucan conversion and chain‐length distribution curves and provide interesting insights into multiple complex and interacting physico‐chemical phenomena involved in enzymatic hydrolysis, including enzyme synergism, substrate accessibility, cellulose chain length distribution and crystallinity, and inhibition of cellulases by soluble sugars. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:1237–1248, 2015     

TLR4‐mediated skin carcinogenesis is dependent on immune and radioresistant cells

Skin cancers are the most commonly diagnosed cancers. Understanding what are the factors contributing to skin tumour development can be instrumental to identify preventive therapies. The myeloid differentiation primary response gene (MyD)88, the downstream adaptor protein of most Toll‐like receptors (TLR), has been shown to be involved in several mouse tumourigenesis models. We show here that TLR4, but not TLR2 or TLR9, is upstream of MyD88 in skin tumourigenesis. TLR4 triggering is not dependent on lipopolysaccharide associated to skin‐colonizing bacteria, but on the high mobility group box‐1 protein (HMGB1), an endogenous ligand of TLR4. HMGB1 is released by necrotic keratinocytes and is required for the recruitment of inflammatory cells and for the initiation of inflammation. The expression of TLR4 on both bone marrow‐derived and radioresistant cells is necessary for carcinogenesis. Consistently, a human tissue microarray analysis showed that melanoma and colon cancer display an over‐expression of TLR4 and its downstream adaptor protein MyD88 within tumours. Together, our results suggest that the initial release of HMGB1 triggers a TLR4‐dependent inflammatory response that leads to tumour development.     

Effect of Organic Selenium Supplementation on Growth, Se Uptake, and Nutrient Utilization in Guinea Pigs

Forty weaned male guinea pigs (Cavia porcellus) of 152.6?±?7.96 g mean body weight were divided into four equal groups and fed a common basal diet comprised of 25% ground cowpea (Vigna unguiculata) hay, 30% ground maize (Zea mays) grain, 22% ground gram (Cicer arietinum) grain, 9.5% deoiled rice (Oryza sativa) bran, 6% soybean (Glycine max) meal, 6% fish meal, 1.5% mineral mixture (without Se), and ascorbic acid at 200 mg/kg to meet their nutrient requirements along with 0, 0.1, 0.2, and 0.3 ppm of organic selenium (Se) in groups I, II, III, and IV, respectively. Experimental feeding lasted for a period of 10 weeks, during which, daily feed intake and weekly body weights were recorded. Intake and digestibility of dry matter, organic matter, ether extract, crude fiber, and nitrogen-free extract as well as uptake of calcium and phosphorus were similar (P?>?0.05) among the four groups. Feed:gain ratio was also similar (P?>?0.05) in the four groups. However, digestibility of crude protein was significantly (P?<?0.001) higher in group II supplemented with 0.1 ppm organic Se as compared to other three group. Intake and absorption of Se was significantly (P?<?0.001) higher in all the Se supplemented groups as compared to control group. Average daily gain (ADG) was significantly (P?<?0.05) higher in group II (3.16 g/day) and III (3.38 g/day) as compared to group I (2.88 g/day). However, ADG in group IV (supplemented 0.3 ppm organic Se) was significantly (P?<?0.05) lower (2.83 g/day) than group II and III, but comparable (P?>?0.05) to group I. Findings of the present experiment suggests that Se requirements of guinea pigs are ≥0.2 ppm, as supplementation of 0.1 ppm organic Se in the diet (having 0.1 ppm Se) not only enhanced their growth rate but also improved the protein utilization.     

A coupled sharp-interface immersed boundary-finite-element method for flow-structure interaction with application to human phonation

A new flow-structure interaction method is presented, which couples a sharp-interface immersed boundary method flow solver with a finite-element method based solid dynamics solver. The coupled method provides robust and high-fidelity solution for complex flow-structure interaction (FSI) problems such as those involving three-dimensional flow and viscoelastic solids. The FSI solver is used to simulate flow-induced vibrations of the vocal folds during phonation. Both two- and three-dimensional models have been examined and qualitative, as well as quantitative comparisons, have been made with established results in order to validate the solver. The solver is used to study the onset of phonation in a two-dimensional laryngeal model and the dynamics of the glottal jet in a three-dimensional model and results from these studies are also presented.     

CHOBIMALT: a cholesterol-based detergent

Cholesterol and its hemisuccinate and sulfate derivatives are widely used in studies of purified membrane proteins but are difficult to solubilize in aqueous solution, even in the presence of detergent micelles. Other cholesterol derivatives do not form conventional micelles and lead to viscous solutions. To address these problems, a cholesterol-based detergent, CHOBIMALT, has been synthesized and characterized. At concentrations above 3?4 μM, CHOBIMALT forms micelles without the need for elevated temperatures or sonic disruption. Diffusion and fluorescence measurements indicated that CHOBIMALT micelles are large (210±30 kDa). The ability to solubilize a functional membrane protein was explored using a G-protein coupled receptor, the human kappa opioid receptor type 1 (hKOR1). While CHOBIMALT alone was not found to be effective as a surfactant for membrane extraction, when added to classical detergent micelles CHOBIMALT was observed to dramatically enhance the thermal stability of solubilized hKOR1.     

Cloning, sequence analysis and crystal structure determination of a miraculin-like protein from Murraya koenigii

Earlier, the purification of a 21.4 kDa protein with trypsin inhibitory activity from seeds of Murraya koenigii has been reported. The present study, based on the amino acid sequence deduced from both cDNA and genomic DNA, establishes it to be a miraculin-like protein and provides crystal structure at 2.9 Å resolution. The mature protein consists of 190 amino acid residues with seven cysteines arranged in three disulfide bridges. The amino acid sequence showed maximum homology and formed a distinct cluster with miraculin-like proteins, a soybean Kunitz super family member, in phylogenetic analyses. The major differences in sequence were observed at primary and secondary specificity sites in the reactive loop when compared to classical Kunitz family members. The crystal structure analysis showed that the protein is made of twelve antiparallel β-strands, loops connecting β-strands and two short helices. Despite similar overall fold, it showed significant differences from classical Kunitz trypsin inhibitors.     

Androgen receptor status predicts response to chemotherapy, not risk of breast cancer in Indian women

Voltage gated potassium channels have been extensively studied in relation to cancer. In this review, we will focus on the role of two potassium channels, Ether à-go-go (Eag), Human ether à-go-go related gene (HERG), in cancer and their potential therapeutic utility in the treatment of cancer. Eag and HERG are expressed in cancers of various organs and have been implicated in cell cycle progression and proliferation of cancer cells. Inhibition of these channels has been shown to reduce proliferation both in vitro and vivo studies identifying potassium channel modulators as putative inhibitors of tumour progression. Eag channels in view of their restricted expression in normal tissue may emerge as novel tumour biomarkers.     

Post-Stroke Inhibition of Induced NADPH Oxidase Type 4 Prevents Oxidative Stress and Neurodegeneration

Ischemic stroke is the second leading cause of death worldwide. Only one moderately effective therapy exists, albeit with contraindications that exclude 90% of the patients. This medical need contrasts with a high failure rate of more than 1,000 pre-clinical drug candidates for stroke therapies. Thus, there is a need for translatable mechanisms of neuroprotection and more rigid thresholds of relevance in pre-clinical stroke models. One such candidate mechanism is oxidative stress. However, antioxidant approaches have failed in clinical trials, and the significant sources of oxidative stress in stroke are unknown. We here identify NADPH oxidase type 4 (NOX4) as a major source of oxidative stress and an effective therapeutic target in acute stroke. Upon ischemia, NOX4 was induced in human and mouse brain. Mice deficient in NOX4 (Nox4 ^−/−) of either sex, but not those deficient for NOX1 or NOX2, were largely protected from oxidative stress, blood-brain-barrier leakage, and neuronal apoptosis, after both transient and permanent cerebral ischemia. This effect was independent of age, as elderly mice were equally protected. Restoration of oxidative stress reversed the stroke-protective phenotype in Nox4 ^−/− mice. Application of the only validated low-molecular-weight pharmacological NADPH oxidase inhibitor, VAS2870, several hours after ischemia was as protective as deleting NOX4. The extent of neuroprotection was exceptional, resulting in significantly improved long-term neurological functions and reduced mortality. NOX4 therefore represents a major source of oxidative stress and novel class of drug target for stroke therapy.