A Step Toward Achieving Peptide Based Therapeutics I. Analysis of the Effect of Olefin Position and Linker Length in Hydrocarbon Stapled Peptides II. Development and Structural Studies of Direct Peptide Inhibitors of Ras

Traditionally the two major classes of drugs, small molecules and biologics, have been extensively used in drug discovery. However, it is difficult to inhibit protein-protein interactions with small molecules, and targets of biologics are limited to extracellular proteins. Considering such limitations, there has been a call for a new therapeutic modality to target ‘undruggable’ proteins. Structurally-stabilized peptide therapeutics has emerged as a new class that could overcome the limitations of the previously established drug classes while retaining their benefits. Among several strategies that have been employed to constrain the conformation of peptides to enhance their pharmacological properties, the all-hydrocarbon stapling system has been particularly successful. This system combines the helix-stabilizing effects of alpha methylation with peptide macrocyclization, which confers improvement not only in helix stabilization but also in protease resistance and cell permeability. The first part of my dissertation focuses on exploring this system further by shortening the established hydrocarbon bridge length as well as moving the position of the olefin along the cross-link, and looking at the effects of these attributes on the helical propensity of the peptides by circular dichroism. In cases where there is no structural information or obvious binding partner that has an alpha-helical component that could be used for rational design of a stapled peptide inhibitor, screening a naïve library of stabilized scaffolds using yeast cell-surface display would be an alternative starting point. Oncogenic K-Ras is one such case; the structures of Ras with its effectors have shown that they interact with Ras via beta-sheet complementation. The second part of my dissertation will describe efforts to develop peptide inhibitors that target Ras by using screening and directed evolution of libraries of an avian pancreatic polypeptide-based scaffold by yeast cell-surface display. We have identified lead peptides that disrupt the binding of effectors to Ras, and the structures of these peptides bound to Ras have been solved and analyzed, leading us to improve the lead peptides further by rational design.Chemistry and Chemical Biolog

Solid fat content and bakery characteristics of interesterified beef tallow-palm mid fraction based margarines

Palm mid fraction (PMF) was interesterified with edible beef tallow (BT) catalyzed using sodium methoxide to investigate the effects on the solid fat content (SFC) of these palmitic rich plastic fats. Interesterified blends crystallize more slowly than BT. Conversely, the crystallization rates of PMF-BT-based interesterification (IE) products were compared with the starting mixture and IE products prepared with non-PMF triglycerides. The SFC PMF-based IE products increased significantly at temperatures between 25 and 40 °C. The SFC profiles became smoother and the products had potential to serve as base oils for preparing specialty fats with a wider range of plasticity. Further exploration of triacylglycerol (TAG) compositional changes revealed that PMF interesterified products had greater saturated/saturated/saturated (S/S/S)-type TAGs compared with soybean oil interesterified products. Moreover, in subsequent evaluations of BT-PMF-based IE fats as a margarine replacement effects in a baked cake model system showed that the material was a suitable functional oil base with acceptable aeration properties and plasticity during baking. Therefore, it is a potential alternative to IE-BT based and traditional IE-BT-palm oil based margarines. The physical-characteristics of bakery products prepared with this fat exhibited improved cake volume with fine structure and clear lifting properties, which affirmed the potential for its application in bakery fats

Chronic Treatment with Squid Phosphatidylserine Activates Glucose Uptake and Ameliorates TMT-Induced Cognitive Deficit in Rats via Activation of Cholinergic Systems

The present study examined the effects of squid phosphatidylserine (Squid-PS) on the learning and memory function and the neural activity in rats with TMT-induced memory deficits. The rats were administered saline or squid derived Squid-PS (Squid-PS 50 mg kg−1, p.o.) daily for 21 days. The cognitive improving efficacy of Squid-PS on the amnesic rats, which was induced by TMT, was investigated by assessing the passive avoidance task and by performing choline acetyltransferase (ChAT) and acetylcholinesterase (AchE) immunohistochemistry. 18F-Fluorodeoxyglucose and performed a positron emission tomography (PET) scan was also performed. In the passive avoidance test, the control group which were injected with TMT showed a markedly lower latency time than the non-treated normal group (P < 0.05). However, treatment of Squid-PS significantly recovered the impairment of memory compared to the control group (P < 0.05). Consistent with the behavioral data, Squid-PS significantly alleviated the loss of ChAT immunoreactive neurons in the hippocampal CA3 compared to that of the control group (P < 0.01). Also, Squid-PS significantly increased the AchE positive neurons in the hippocampal CA1 and CA3. In the PET analysis, Squid-PS treatment increased the glucose uptake more than twofold in the frontal lobe and the hippocampus (P < 0.05, resp.). These results suggest that Squid-PS may be useful for improving the cognitive function via regulation of cholinergic enzyme activity and neural activity

Biorefinery process for protein extraction from oriental mustard (Brassica juncea (L.) Czern.) using ethanol stillage

Large volumes of treated process water are required for protein extraction. Evaporation of this water contributes greatly to the energy consumed in enriching protein products. Thin stillage remaining from ethanol production is available in large volumes and may be suitable for extracting protein rich materials. In this work protein was extracted from ground defatted oriental mustard (Brassica juncea (L.) Czern.) meal using thin stillage. Protein extraction efficiency was studied at pHs between 7.6 and 10.4 and salt concentrations between 3.4 × 10-2 and 1.2 M. The optimum extraction efficiency was pH 10.0 and 1.0 M NaCl. Napin and cruciferin were the most prevalent proteins in the isolate. The isolate exhibited high in vitro digestibility (74.9 ± 0.80%) and lysine content (5.2 ± 0.2 g/100 g of protein). No differences in the efficiency of extraction, SDS-PAGE profile, digestibility, lysine availability, or amino acid composition were observed between protein extracted with thin stillage and that extracted with NaCl solution. The use of thin stillage, in lieu of water, for protein extraction would decrease the energy requirements and waste disposal costs of the protein isolation and biofuel production processes

Tension pneumopericardium after removal of pericardiocentesis drainage catheter

This image showed tension pneumopericardium caused by removing the pericardiocentesis catheter, which was inserted to drain malignant pericardial effusion. Tension pneumopericardium is a rare and potentially fatal event. Mortality from tension pneumopericardium can be as high as 50%. Therefore, it is important to suspect and detect early, if the patient complained of dyspnea after removing the pericardiocentesis drainage cathete

Novel Flaxseed Gum Nanocomposites are Slow Release Iron Supplements

Nanocomposites, based on iron salts and soluble flaxseed gum (FG), were prepared as potential treatments of iron deficiency anemia (IDA). FG was extracted, characterized, and formulated into iron-loading nanocomposites via ion-exchange against FeCl_3, Fe_2(SO_4)_3, FeCl_2, and FeSO_4·7H_2O. FG-iron nanocomposites preparation condition was optimized, and physicochemical properties of the nanocomposites were investigated. In vitro release kinetics of iron in simulated gastric fluid (SGF) was also evaluated. FG heteropolysaccharide, consisting of rhamnose (33.73%), arabinose (24.35%), xylose (14.23%), glucose (4.54%), and galactose (23.15%) monosaccharides, linked together via varieties of glycosidic bonds, was a good recipient for both ferric and ferrous irons under screened conditions (i.e., 80 °C, 2 h, I/G = 1:2). Iron loaded contents in the nanocomposites prepared from FG-FeCl_3, FG-Fe_2(SO_4)_3, FG-FeCl_2, and FG-FeSO_4·7H_2O were 25.51%, 10.36%, 5.83%, and 22.83%, respectively. Iron in these nanocomposites was mostly in a bound state, especially in FG-FeCl_3, due to chelation forming bonds between iron and polysaccharide hydroxyl or carboxyl groups and formed stable polysaccharide-iron crystal network structures. Free iron ions were effectively removed by ethanol treatments. Because of chelation, the nanocomposites delayed iron release in SGF and the release kinetics were consistent with Korsmeyer-Peppas model. This indicates that such complexes might reduce side effects of free iron in human stomach. Altogether, this study indicates that these synthetic FG-iron nanocomposites might be developed as novel iron supplements for iron deficiency, in which FG-FeCl_3 is considered as the best option

Computational analysis of the CB1 carboxyl-terminus in the receptor-G protein complex: Computational Analysis of the CB1 Carboxyl-Terminus

Despite the important role of the carboxyl-terminus (Ct) of the activated brain cannabinoid receptor one (CB1) in the regulation of G protein signaling, a structural understanding of interactions with G proteins is lacking. This is largely due to the highly flexible nature of the CB1 Ct that dynamically adapts its conformation to the presence of G proteins. In the present study, we explored how the CB1 Ct can interact with the G protein by building on our prior modeling of the CB1-Gi complex (Shim J-Y, Ahn KH, Kendall DA. The Journal of Biological Chemistry 2013;288:32449-32465) to incorporate a complete CB1 Ct (Glu416Ct–Leu472Ct). Based upon the structural constraints from NMR studies, we employed ROSETTA to predict tertiary folds, ZDOCK to predict docking orientation, and molecular dynamics (MD) simulations to obtain two distinct plausible models of CB1 Ct in the CB1-Gi complex. The resulting models were consistent with the NMR-determined helical structure (H9) in the middle region of the CB1 Ct. The CB1 Ct directly interacted with both Gα and Gβ and stabilized the receptor at the Gi interface. The results of site-directed mutagenesis studies of Glu416Ct, Asp423Ct, Asp428Ct, and Arg444Ct of CB1 Ct suggested that the CB1 Ct can influence receptor-G protein coupling by stabilizing the receptor at the Gi interface. This research provided, for the first time, models of the CB1 Ct in contact with the G protein