Biotechnology and Bioengineering

The authors present a Zonal Rate Model that is shown to improve upon current mathematical models for the description of protein breakthrough from stacked membrane chromatography. The model implements a virtual radial partitioning of both the membrane stack and external hold up volumes thus accurately describing all nonidealities related to dispersion and flow distribution. This, in turn, allows for a rigorous evaluation of putative protein adsorption models onto an ion exchange membrane chromatography module.

CYP51 and iron-sulfur containing FprD were fused together with designed linker sequences. CYP51-FprD fusion enzymes showed distinct spectral properties of both flavoprotein and CYP. CYP51-FprD F1 and F2 in recombinant E. coli BL21(DE3) catalyzed demethylation of lanosterol more efficiently, with k_cat/K_m values of 96.91 and 105.79 nmol/min⁻¹/nmol, respectively, which are about 35-fold higher compared to those of CYP51 and FprD alone.

Elemental sulfur was used as an electron donor to reduce perchlorate to harmless products in a pilot scale packed bed bioreactor at the Massachusetts Military Reservation. Issues of scale-up, groundwater deoxygenation and the effect of the co-contaminants, nitrate and RDX, are discussed. The microbial community structure was investigated using DGGE community fingerprint analysis, and functional gene amplification, identification and quantification. Molecular biology results provided insights into bioreactor performance that could not be gained through examination of the water quality data.

A spectral deconvolution method was developed to rapidly and quantitatively determine syringyl to guaiacyl ratios of plant materials using FT-Raman spectroscopy. The method was demonstrated to be a valid method to measure a wide range of S/G ratios derived from different plant species, evaluate genetic modifications, compare ecotypes, and assess plants at different developmental stages.

A mechanistic model for the enzymatic saccharification of cellulose has been developed. In part I, the mathematical description of the polymeric cellulose and its representation as a solid substrate with limited accessibility is presented. The distinct modes of action of an endoglucanase (EGI) and an exoglucanase (CBHI), which are responsible for cellulose deconstruction, are also accounted for in the model. Model simulations illustrate the time evolution of cellulose population distributions that result from enzymatic digestion.

A mechanistic model for the enzymatic saccharification of cellulose has been developed. In part II, the kinetic model from part I is expanded to include solution-phase kinetics, an innovative product-inhibition scheme, and sterically hindered enzyme adsorption, based on current structural knowledge of the component enzymes. The model was used to explore several potential rate-limiting phenomena (e.g. accessibility, inhibition, degree of polymerization). Preliminary simulation results indicate that the accessibility of the substrate to the enzymes is a key rate-determining factor.

The present investigation is aimed towards the production of ethanol from complex lignocellulosic biomass. Thermophilic bacterium, isolated from a hot spring in Iceland, was used for production of ethanol and hydrogen with the main aim to investigate sustainable biofuel production. The pros and cons of using thermophiles for biofuel production were described and the effect of various factors to maximize ethanol yields were investigated.

The effects of the directionality of pulsatile WSS on endothelial cell proliferation and morphology were investigated for cells grown in a Petri dish orbiting on a shaker platform. Since WSS on the bottom of the dish is two-dimensional, a new directional oscillatory shear index (DOSI) was developed to quantify the directionality of oscillating shear. Statistical analyses of the individual and interacting effects of multiple factors (DOSI, shear magnitudes, and orbital speeds) showed that DOSI significantly affected all the responses, indicating that directionality is an important determinant of cellular responses.

By combining adaptive evolution strategy and high-density fermentation mode, the authors successfully evolved a Clostridium tyrobutyricum mutant with significantly improved butyric acid volumetric productivity and enhanced glucose consumption rate.

The solubility of cytochrome c in various ionic liquids (ILs) has been analyzed. The solubility showed a good relationship with polarity parameters of ILs. The dissolved cytochrome c was found to keep its redox activity in some polar ILs. The redox response of the dissolved cytochrome c was detected in 1-allyl-3-methylimidazolium chloride up to 140 ^°C.

The addition of copper ion to a mammalian cell culture harvest results in target enzyme binding to an uncharged metal chelate column. 5mM copper is optimal for target enzyme recovery.

The removal of analog impurities presents a major challenge in the purification of high-value biochemicals (such as those derived from fermentation or herbs) within acceptable cost and time constraints. Ganoderic acid T (GA-T), an antitumor drug candidate, is very difficult to purify from the mycelia of medicinal mushroom Ganoderma lucidum due to co-purifying, analog impurities. A novel pretreatment process with three consecutive chemical conversion steps, namely hydrolysis–acetylation–hydrolysis, was developed to convert two key analog impurities to GA-T.

For ¹³C metabolic flux analysis applied to microbial central carbon metabolism, this article demonstrates that the precision of estimation is improved with tandem mass spectrometry. Specifically, the fragmentation induced in a collision cell provides positional information on ¹³C enrichment which is missed when analyzing only whole intermediates of central metabolism. Additionally, a special mass spectrometry acquisition technique is presented to enable efficient measurement of a large set of intact metabolites and fragments thereof.

The image depicts a schematic representation of the cell cycle considered in this work.

Simulated temporal evolution of the number percentages of cells falling into the different cell cycle phases ( with P = G1, S, G2/M) and the normalized total counts are presented.

Kidney epithelial cells reclaim protein through receptor mediated endocytosis. Shear stress alters kidney epithelial cell response to protein exposure. The image shows uptake of fluorescently labeled albumin by opossum kidney epithelial cells exposed to fluid flow.

Large eddy simulation of mechanical mixing in anaerobic digesters is conducted using Smagorinsky-Lilly, wall-adapting local eddy-viscosity, and kinetic energy transport models, respectively, in which the impeller rotation is characterized by the sliding mesh method. The simulation results indicate that the kinetic energy transport model performs the best in terms of predicting the impeller power and flow numbers, and that a large eddy simulation performs better than a Reynolds-averaged Navier-Stokes simulation in the study of turbulent flow.

Controlling the differentiated phenotype of embryonic stem cell-derived cells is critical to the development of stem cell-based therapies. This work explores the use of controlled topographical cues from synthetic scaffolds and coculture with fibroblasts to influence the phenotype of embryonic stem cell-derived cardiomyocytes for use in cardiac tissue engineering applications.

This work describes a novel strategy to deliver therapeutic molecules into macrophages, taking advantage of CD163 receptor-mediated endocytosis of hemoglobin-haptoglobin complexes. The drug delivery system described here consists of hemoglobin (Hb) surface decorated liposomes that can encapsulate any therapeutic molecule of interest. The results of this study clearly demonstrate that this delivery system is specific towards macrophages and demonstrates the feasibility of using this approach in targeted drug delivery.

An in vitro co-culture system has been developed to investigate neurite extension as a function of hydrogel design parameters using gene delivery. Accessory cells (blue) migrate through the hydrogel and access the entrapped DNA (red, left) that encodes for nerve growth factor (NGF). The sustained expression of NGF supports neurite outgrowth (red, right) of the co-encapsulated DRG explants. The hydrogel design that effectively balances the requirements for nerve growth and gene delivery maximized neurite outgrowth within hydrogels.

The use of membrane filtration for removing protein aggregates in protein solutions may result in the additional effect of aggregate fragmentation. Fragmented aggregates, smaller than the membrane pore size, may be generated and passed through the membrane, diminishing the net benefit of filtration for aggregate removal. The presence of protein aggregates in a solution can significantly accelerate the kinetics of amyloid formation, which is associated with diseases such as Alzheimer's.

A robust model developed for predicting toxicity in E. coli is presented in this work. The predictor based on experimental toxicity values can be used in many applications from metabolic engineering to drug design, representing a valuable tool for synthetic biology and drug discovery.