Nanotubular structures developed from whey-based α-lactalbumin fractions for food applications

Whey proteins have high nutritional value providing use in dietary purposes and improvement of technological properties in processed foods. Functionality of the whey-based α-lactalbumin (α-La) may be increased when assembled in the form of nanotubes, promising novel potential applications subject to investigation. The purpose of this study was to extract highly pure α-La from whey protein isolate (WPI) and whey powder (WP) and to construct protein nanotubes from them for industrial applications. For protein fractionation, WPI was directly fed to chromatography, however, WP was first subjected to membrane filtration and the retentate fraction, whey protein concentrate (WPC), was obtained and then used for chromatographic separation. α-La and, additionally β-Lg, were purified at the same batches with the purities in the range of 95%-99%. After enzymatic hydrolysis, WPI-based α-La produced chain-like and long nanotubules with ∼20 nm width while WPC-based α-La produced thinner, miscellaneous, and fibril-like nanostructures by self-assembly. Raman and FT-IR spectroscopies revealed that α-La fractions, obtained from both sources and the nanostructures, developed using both fractions have some structural differences due to conformation of secondary structure elements. Nanotube formation induced gelation and nanotubular gel network entrapped a colorant uniformly with a transparent appearance. Dairy-based α-La protein nanotubules could be served as alternative gelling agents and the carriers of natural colorants in various food processes.Scientific and Technological Research Council of Turkey(109O866); Izmir Institute of Technology (IYTE03

Additional file 3: Figure S2. of SPEN, a new player in primary cilia formation and cell migration in breast cancer

SPEF2 silencing in T47D-SPEN cells inhibits cellular migration. (a) Western blot analysis of SPEF2 protein levels in T47D-SPEN cells treated with a CTL siRNA (siCTL) or two different SPEF2 siRNAs (siSPEF2#1 and #2) and T47D-CTL cells treated with a CTL siRNA (siCTL; two independent lysates). (b) Effect of SPEF2 knockdown on the migration of T47D SPEN cells was evaluated by performing Transwell migration assays. Each error bar represents the mean and SEM of three independent experiments performed in duplicate. *p < 0.05, **p < 0.01. (c) Representative images of migrated cells fixed and stained with crystal violet after 72 h. (PDF 456 kb

Additional file 2: Figure S1. of SPEN, a new player in primary cilia formation and cell migration in breast cancer

MCF10A cells grown with or without cholera toxin display important phenotypic and functional differences. (a and b) Representative pictures of MCF10A cells grown in the absence (a) or presence (b) of cholera toxin. (c and d) Representative images of primary cilia in MCF10A cells grown in the absence (c) or presence (d) of cholera toxin. Arrows point to primary cilia (scale bar = 5 μm). (e) Cell cycle analyses performed with MCF10A cells grown in the absence and presence of cholera toxin. Bar graph represents the mean percentage of cells (±SEM) in each phase of the cell cycle in three independent experiments. (f) Growth curve of MCF10A cells grown in the presence or absence of cholera toxin. Data points represent the mean fluorescence values (±SEM) of three experiments performed in quadruplicates. (PDF 9130 kb

Conformation-Based Restrictions and Scaffold Replacements in the Design of Hepatitis C Virus Polymerase Inhibitors: Discovery of Deleobuvir (BI 207127)

Conformational restrictions of flexible torsion angles were used to guide the identification of new chemotypes of HCV NS5B inhibitors. Sites for rigidification were based on an acquired conformational understanding of compound binding requirements and the roles of substituents in the free and bound states. Chemical bioisosteres of amide bonds were explored to improve cell-based potency. Examples are shown, including the design concept that led to the discovery of the phase III clinical candidate deleobuvir (BI 207127). The structure-based strategies employed have general utility in drug design

Discovery of the First Thumb Pocket 1 NS5B Polymerase Inhibitor (BILB 1941) with Demonstrated Antiviral Activity in Patients Chronically Infected with Genotype 1 Hepatitis C Virus (HCV)

Combinations of direct acting antivirals (DAAs) that have the potential to suppress emergence of resistant virus and that can be used in interferon-sparing regimens represent a preferred option for the treatment of chronic HCV infection. We have discovered allosteric (thumb pocket 1) non-nucleoside inhibitors of HCV NS5B polymerase that inhibit replication in replicon systems. Herein, we report the late-stage optimization of indole-based inhibitors, which began with the identification of a metabolic liability common to many previously reported inhibitors in this series. By use of parallel synthesis techniques, a sparse matrix of inhibitors was generated that provided a collection of inhibitors satisfying potency criteria and displaying improved in vitro ADME profiles. “Cassette” screening for oral absorption in rat provided a short list of potential development candidates. Further evaluation led to the discovery of the first thumb pocket 1 NS5B inhibitor (BILB 1941) that demonstrated antiviral activity in patients chronically infected with genotype 1 HCV

Minimizing the Contribution of Enterohepatic Recirculation to Clearance in Rat for the NCINI Class of Inhibitors of HIV

A scaffold replacement approach was used to identifying the pyridine series of noncatalytic site integrase inhibitors. These molecules bind with higher affinity to a tetrameric form compared to a dimeric form of integrase. Optimization of the C6 and C4 positions revealed that viruses harboring T124 or A124 amino acid substitutions are highly susceptible to these inhibitors, but viruses having the N124 amino acid substitution are about 100-fold less susceptible. Compound <b>20</b> had EC₅₀ values <10 nM against viruses having T124 or A124 substitutions in IN and >800 nM in viruses having N124 substitions. Compound <b>20</b> had an excellent in vitro ADME profile and demonstrated reduced contribution of biliary excretion to in vivo clearance compared to BI 224436, the lead compound from the quinoline series of NCINIs

Aligning Potency and Pharmacokinetic Properties for Pyridine-Based NCINIs

Optimization of pyridine-based noncatalytic site integrase inhibitors (NCINIs) based on compound <b>2</b> has led to the discovery of molecules capable of inhibiting virus harboring N124 variants of HIV integrase (IN) while maintaining minimal contribution of enterohepatic recirculation to clearance in rat. Structure–activity relationships at the C6 position established chemical space where the extent of enterohepatic recirculation in the rat is minimized. Desymmetrization of the C4 substituent allowed for potency optimization against virus having the N124 variant of integrase. Combination of these lessons led to the discovery of compound <b>20</b>, having balanced serum-shifted antiviral potency and minimized excretion in to the biliary tract in rat, potentially representing a clinically viable starting point for a new treatment option for individuals infected with HIV