Mechanisms of Enhanced Catalysis in Enzyme–DNA Nanostructures Revealed through Molecular Simulations and Experimental Analysis

Understanding and controlling the molecular interactions between enzyme substrates and DNA nanostructures has important implications in the advancement of enzyme–DNA technologies as solutions in biocatalysis. Such hybrid nanostructures can be used to create enzyme systems with enhanced catalysis by controlling the local chemical and physical environments and the spatial organization of enzymes. Here we have used molecular simulations with corresponding experiments to describe a mechanism of enhanced catalysis due to locally increased substrate concentrations. With a series of DNA nanostructures conjugated to horseradish peroxidase, we show that binding interactions between substrates and the DNA structures can increase local substrate concentrations. Increased local substrate concentrations in HRP(DNA) nanostructures resulted in 2.9‐ and 2.4‐fold decreases in the apparent Michaelis constants of tetramethylbenzidine and 4‐aminophenol, substrates of HRP with tunable binding interactions to DNA nanostructures with dissociation constants in the micromolar range. Molecular simulations and kinetic analysis also revealed that increased local substrate concentrations enhanced the rates of substrate association. Identification of the mechanism of increased local concentration of substrates in close proximity to enzymes and their active sites adds to our understanding of nanostructured biocatalysis from which we can develop guidelines for enhancing catalysis in rationally designed systems.     

Thermal conductivity of the ionic liquid [HMIm][Tf2N] with compressed carbon dioxide

The liquid thermal conductivity of the ionic liquid (IL), 1-hexyl-3-methyl-imidazolium bis(trifluoromethylsulfonyl)amide ([HMIm][Tf₂N]), saturated with compressed vapor and supercritical carbon dioxide was measured over three isotherms (298.15, 323.15, and 348.15 K) and pressures up to approximately 20 MPa using a transient hot-wire technique. Pure [HMIm][Tf₂N] thermal conductivity was also measured over a temperature range of 293.15–353.15 K at ambient pressure and with hydrostatic pressure to approximately 20 MPa. Literature vapor–liquid equilibrium data were used to predict the liquid CO₂ composition at the conditions investigated. Initially, the liquid thermal conductivity slightly decreased with pressure/composition of CO₂ followed by a gradual increase that is mainly attributed to hydrostatic pressure effects. Simple composition-based mixing rules for mixture properties are not qualitatively nor quantitatively accurate. These data could be used to engineer heat transfer equipment required for a variety of proposed IL applications in CO₂ capture, absorption refrigeration, biphasic CO₂/IL reaction platforms, etc.     

Phospholipid Cyclosporine Prodrugs Targeted at Inflammatory Bowel Disease (IBD) Treatment: Design,Synthesis, and in Vitro Validation

Novel phospholipid (PL)-cyclosporine conjugates were prepared and studied as potential prodrugs for inflammatory bowel disease (IBD). Our approach relies on phospholipase A₂ (PLA₂), which is overexpressed in the inflamed intestinal tissues, as the prodrug activator to potentially release cyclosporine at the site of inflammation. PL-cyclosporine prodrug conjugates with methylene linkers of various lengths between the sn-2 position of the PL and cyclosporine were synthesized and evaluated for in vitro activation. Surprisingly, despite previous work indicating that conjugates with six methylene linkers between the lipid and drug would suffer rapid enzymatic hydrolysis, with cyclosporine this was not observed. However, compounds with longer linkers (n=10, 12 methylene units) display complete release of the drug by PLA₂-catalyzed hydrolysis, thus demonstrating the importance and profound impact of structural fine-tuning. This study represents a proof-of-concept for our hypothesis and a first step towards a truly targeted IBD treatment with cyclosporine that could be administered throughout the GI tract.     

Biophysical Modulation of Mesenchymal Stem Cell Differentiation in the Context of Skeletal Repair

A prominent feature of the skeleton is its ability to remodel in response to biophysical stimuli and to repair under varied biophysical conditions. This allows the skeleton considerable adaptation to meet its physiological roles of stability and movement. Skeletal cells and their mesenchymal precursors exist in a native environment rich with biophysical signals, and they sense and respond to those signals to meet organismal demands of the skeleton. While mechanical strain is the most recognized of the skeletal biophysical stimuli, signaling phenomena also include fluid flow, hydrostatic pressure, shear stress, and ion-movement-related electrokinetic phenomena including, prominently, streaming potentials. Because of the complex interactions of these electromechanical signals, it is difficult to isolate the significance of each. The application of external electrical and electromagnetic fields allows an exploration of the effects of these stimuli on cell differentiation and extra-cellular matrix formation in the absence of mechanical strain. This review takes a distinctly translational approach to mechanistic and preclinical studies of differentiation and skeletal lineage commitment of mesenchymal cells under biophysical stimulation. In vitro studies facilitate the examination of isolated cellular responses while in vivo studies permit the observation of cell differentiation and extracellular matrix synthesis.     

碳水化合物——蛋白质饮料对运动耐力的影响

人在长时间运动后的疲劳和体内存储的肌糖原的消耗密切相关,因此运动后糖原合成的速度是短时间内运动恢复的关键因素。接下来的实验有效地研究了运动后补充的营养成分对于再次运动时耐力表现的影响。     

Recent advances refining galactooligosaccharide production from lactose

The production of prebiotic galactooligosaccharides (GOS) by the β-galactosidase catalysed conversion of lactose has become commercially important. Yet it remains a challenge to sufficiently understand the structure and activity of β-galactosidase, to increase the efficiency of transgalactosylation and GOS production and to improve the quality of GOS products in a rational way. This review covers the broad but related aspects of GOS synthesis including: the structure and reaction mechanism of β-galactosidase, factors effecting yield and productivity of GOS synthesis systems, the structure of GOS products, models for the kinetics of GOS synthesis and reactor configurations for GOS synthesis. It aims to couple recent discoveries with established knowledge to enhance understanding of the complex biochemistry of GOS synthesis.     

Development of Undergraduate Students’ Professional Skills

The development of engineering students’ professional skills has gained considerable national attention from Accreditation Board for Engineering and Technology, the National Academy of Engineering, ASCE, and other constituents. There is little debate that these professional skills are necessary. Engineering programs have tried many approaches to develop these skills in the undergraduate programs. Colorado State University (CSU) has developed a new approach modeled on the type of professional development that occurs in the professional environment. This new Professional Learning Institute (PLI) provides students with a broad array of workshops, presentations, and experiential opportunities addressing the areas of cross cultural communication and teamwork, innovation, leadership, ethics, and public service. This program introduces students to the concept of professional development through required extracurricular activities, includes minimum requirements along with requirements to earn certificates in specialty areas for motivated students. The majority of offerings in the PLI are presented by leaders from the engineering profession who have teamed with CSU to provide high quality programs for our students.