Structure and functionality of almond proteins as a function of pH

Molecular mechanism for stabilization of a mutant α-galactosidase A involving M51I amino acid substitution by imino sugars

Theoretical study of chlordecone and surface groups interaction in an activated carbon model under acidic and neutral conditions

The use of surfactants as additives was demonstrated for the first time in capillary isoelectric focusing (CIEF) to dynamically modify the surfaces of bare fused silica capillaries. These surfactants were zwitterionic sulfobetaines: dodecyldimethyl (3-sulfopropyl) ammonium hydroxide (C12N3SO3), hexadecyldimethyl (3-sulfopropyl) ammonium hydroxide (C16N3SO3) and coco (amidopropyl)hydroxyldimethylsulfobetaine (Rewoteric AM CAS U). They were added directly to the protein-ampholyte mixture, and remained in the capillary during isoelectric focusing and mobilization. The C16N3SO3 and CAS U coatings were shown effective in CEF. Separation of seven IEF protein standards was obtained, with significantly improved resolution compared to that from an uncoated silica capillary. The effect of these surfactants on the electroosmotic flow (EOF) in CIEF was determined. CAS U was effective in suppressing the EOF at neutral and alkaline pH conditions, C16N3SO3 was effective in suppressing EOF at acidic and neutral pH conditions. C12N3SO3 however had little effect on the EOF. The pH gradients formed inside these surfactant coated capillaries were recta-linear at pH 6 to 9 (R2 approximately equal to 0.99). Reproducibility of migration time and peak area was determined. For all three coatings, the migration time standard deviations were less than 1.6 min, and the relative standard deviations of area were below 10%. The protein recovery in the CAS U-modified capillary was quantitative or near-quantitative for five of the seven proteins studied.

Plant-based diets have become popular in the past 10 years, with approximately 11% of Americans self-identifying as vegan or vegetarian, and many others trying to reduce meat consumption. Due to this increasing interest, the plant-based food market has exploded with several novel innovative products serving as alternatives to animal-based products. One such example is almond protein powder, a fairly new protein supplement created as an alternative to whey protein. A number of studies have compared animal-based protein supplementation, such as whey to plant-based supplementation, such as soy, on muscle protein synthesis and skeletal muscle preservation. Due to the novelty of almond protein, little is known regarding how well the protein supplement performs in the body. The effects of both almond and whey-based protein beverage on nitrogen balance, body composition, and hydration in the body were investigated in the work presented herein. Twenty female students aged 20-25 were randomly assigned to consume either an almond or whey-based protein drink twice daily for 7 days. A 24-hour urine collection was performed at baseline and endpoint of the 7-day treatment period, and nitrogen balance was assessed. The effects of supplementation on nitrogen balance in almond and whey protein were equally capable of increasing significantly the N balance from 8.58g to 11.66g (p =0.05), indicating that almond protein powder may be a functional plant-based replacement to whey protein powder, and may be of interest in future research regarding muscle mass and body composition changes.

Conformational study of globulin from rice ( Oryza sativa) seeds by Fourier-transform infrared spectroscopy

FT-IR spectroscopy has been applied to study the secondary structure of the major coat protein of Pf1 and M13 as present in the phage and reconstituted in DOPG and mixed DOPC/DOPG (4/1) bilayers. Infrared absorbance spectra of the samples were examined in dehydrated films and in suspensions of D2O and H2O. The secondary structure of the coat protein is investigated by second-derivative analysis, Fourier self-deconvolution, and curve fitting of the infrared bands in the amide I region (1600-1700 cm-1). It is found that, in dehydrated films of Pf1 and M13 phage, the amide I region contains three bands located at about 1633, 1657, and 1683 cm-1, that are assigned to hydrogen-bonded turn, alpha-helix/random coil, and non-hydrogen-bonded turn, respectively. From a comparison of the infrared spectra in dehydrated film with those in aqueous suspension, the percentages of secondary structure were found with an accuracy of about +/- 5%. For the coat protein of Pf1 phage, the FT-IR quantification gives 69% alpha-helix conformation, 19% turn structure, and 12% random coil structure. For Pf1 coat protein in the membrane-embedded state, the amount of alpha-helix is 57%, whereas 42% is in a turn structure and 1% in a random coil structure. The same assignment strategy was used for the analysis of the data obtained for M13 coat protein reconstitution into phospholipid systems. For M13 coat protein in the phage, this gives 75% alpha-helix conformation, 21% turn structure, and 4% random coil structure.(ABSTRACT TRUNCATED AT 250 WORDS)

Background and ObjectivesPlant protein products are now considered sustainable substitutes for animal proteins in new product formulations, and various methods such as ultrasound have been used to improve their properties. The aim of this study was to investigate the physicochemical and functional properties of grass pea (Lathyrus sativus L.) protein isolate was extracted, as a promising source of plant‐based protein, using conventional (CP), pre‐ultrasonic (PUP) and ultrasound‐assisted (UAP) treatments.FindingsExtraction yield, protein content, and protein yield of ultrasound treatments were increased. Ultrasound treatments improved the functional properties of pea protein. Thermal and surface properties were also altered due to the conformational changes in protein molecules during sonication. According to sodium dodecyl sulfate‐polyacrylamide gel electrophoresis, there were no differences in protein fractions of different samples in terms of molecular weight, and protein fractions between 23 and 100 kDa were observed. However, Fourier transform infrared spectroscopy results revealed changes in different treatments suggesting that sonication changed protein secondary structures. Correspondingly, optical and fluorescence microscopy images of protein solutions and foams revealed changes in the structure of proteins and CP presented larger protein aggregates and larger foam bubble sizes than PUP and UAP.ConclusionsUltrasound treatment could increase extraction and improve the functionality of proteins which was attributed to changes in the secondary structure of proteins.Significance and NoveltyGrass pea protein isolate can be considered a novel source of valuable protein and ultrasound treatment could enhance the functionality of the protein increasing potential applications of this ingredient in the food industry.

Phenolic compounds can undergo auto-oxidation, especially at alkaline pH, forming reactive o -quinones that bind covalently to proteins and may affect the protein structure, solubility, and functional properties. Yet, it is still unclear how the phenolic compounds' structure and properties affect their reaction with proteins, and how they influence the resulting changes in protein structure and functionality. Therefore, the model protein β-lactoglobulin (BLG) was incubated with ten common phenolic compounds at pH 8.5 for 24 h at a phenolic-to-protein molar ratio of 5:1. RP-HPLC was used to screen for covalent modifications. Protein structural changes were investigated using MALDI-TOF-MS, OPA and Ellman's assays, ATR-FTIR, tryptophan fluorescence quenching, SDS-PAGE, and SEC. Protein functionality changes were determined by oil droplet size measurement after emulsion formation via high pressure homogenization. Only phenolic compounds with a di or-trihydroxybenzene moiety resulted in noteworthy BLG modification (>40 %), with an average of one phenolic compound bound per protein molecule, primarily on the thiol groups of the cysteine residues. Modification decreased the protein's α-helices and shifted the intramolecular β-sheets to higher wavelengths. Protein modification resulted in the formation of smaller oil droplets (from 1.38 to 2 μm) at low homogenization pressure. A positive relationship was found between the presence of a carboxyl group in the phenolic compounds, the unfolding of the protein's tertiary structure (R 2 = 0.95), and smaller oil droplet size. The insights from this study support the selection of phenolic compounds for targeted protein modifications or for avoiding undesired protein modifications. • Only phenolics with a catechol or pyrogallol moiety modify proteins at alkaline pH • At pH 8.5, phenolic compounds primarily bind at the free thiol groups of cysteines • Modification results in a decrease and shift in intramolecular β-sheets • Phenolic compounds with a carboxyl group induced protein tertiary structure changes • Protein tertiary structure changes lead to a decrease in emulsion droplet size

Cathepsin L plays physiological and pathological roles in immune responses, cancer, metamorphosis, and oogenesis in several species. However, the function of Cathepsin L in medaka ovaries remains unclear. Therefore, here, we examined the physiological functions of Cathepsin L in the medaka ovaries. Cathepsin L mRNA transcripts and proteins were found to be constitutively expressed in the ovaries of Oryzias latipes over a 24-h spawning cycle. Expression was localized within the oocyte cytoplasm of growing follicles and the follicle layer of preovulatory and postovulatory follicles. Moreover, the active form of Cathepsin L was highly expressed in the follicle layer of periovulatory follicles and the ovaries 2-6h after ovulation. Recombinant Cathepsin L was activated under acidic conditions and exhibited enzymatic activity in acidic and neutral pH conditions. However, extracellular matrix proteins were degraded by recombinant Cathepsin L under acidic, not neutral pH conditions. Cathepsin L was secreted from preovulatory follicles, while active recombinant Cathepsin L was detected in the conditioned medium of a medaka cell line, OLHNI-2. Mechanistically, recombinant Cathepsin L activates recombinant urokinase-type plasminogen activator-1, which is expressed within the follicle layers post-ovulation. Meanwhile, the treatment of medakas with an E-64 or anti-Cathepsin L antibody effectively blocked follicular layer degeneration and degradation after ovulation, whereas in vitro ovulation was not inhibited by either. Collectively, the findings of this study indicate that although Cathepsin L does not impact ovulation in medakas, it contributes to the degeneration and degradation of the follicle layers following ovulation via activation of urokinase-type plasminogen activator-1, and not via the degradation of extracellular matrix proteins.

Silk from Indian paper wasp: Structure prediction and secondary conformational analysis

Degradation of dried Ti-peroxy gels made from metallic titanium and hydrogen peroxide

HbPresbyterian (beta 108Asn --> Lys, HbP) contains an additional positive charge (per alpha beta dimer) in the middle of the central cavity and exhibits a lower oxygen affinity than wild-type HbA in the presence of chloride. However, very little is known about the molecular origins of its altered functional properties. In this study, we have focused on the beta beta cleft of the Hb tetramer. Recently, we developed an approach for quantifying the ligand binding affinity to the beta-end of the Hb central cavity using fluorescent analogues of the natural allosteric effector 2, 3-diphosphoglycerate (DPG) [Gottfried, D. S., et al. (1997) J. Biol. Chem. 272, 1571-1578]. Time-correlated single-photon counting fluorescence lifetime studies were used to assess the binding of pyrenetetrasulfonate to both HbA and HbP in the deoxy and CO ligation states under acidic and neutral pH conditions. Both the native and mutant proteins bind the probe at a weak binding site and a strong binding site; in all cases, the binding to HbP was stronger than to HbA. The most striking finding was that for HbA the binding affinity varies as follows: deoxy (pH 6.35) > deoxy (pH 7.20) > CO (pH 6.35); however, the binding to HbP is independent of ligation or pH. The mutant oxy protein also hydrolyzes p-nitrophenyl acetate, through a reversible acyl-imidazole pathway linked to the His residues of the beta beta cleft, at a considerably higher rate than does HbA. This implies a perturbation of the microenvironment of these residues at the DPG binding pocket. Structural consequences due to the presence of the new positive charge in the middle of the central cavity have been transmitted to the beta beta cleft of the protein, even in its liganded conformation. This is consistent with a newly described quaternary state (B) for liganded HbPresbyterian and an associated change in the allosteric control mechanism.

A fluorophore-anchor dye based on the 4-amino-1,8-naphthalimide chromophore was designed and characterised with respect to its ability of forming supramolecular host–guest complexes with cucurbit[6]uril (CB6) and cucurbit[7]uril (CB7) hosts. It was found that CB6 encapsulates the anchor of the dye with a high binding constant [K = (1.1–1.4) × 107 M− 1], which was independent of acidic or neutral pH conditions. The 1:1 binding was accompanied by fluorescence quenching (ca. 20%) and a bathochromic shift (Δλ = +22 nm) of the charge-transfer absorption band of the dye. This is indicative of the occurrence of hydrogen bonding between a carbonyl-lined host portal and the aromatic NH group of the guest dye. In comparison, CB7 encapsulated the dye much less efficiently (K = 4.8 × 104 M− 1) and showed a significant fluorescence enhancement (Φf = 0.15 versus 0.52 for free and CB7-complexed dye at pH 4, respectively). The occurrence of hydrogen bonding with the aromatic NH was much less evident for CB7 as judged by the only minor bathochromic shift (Δλ = +4 nm) of the charge-transfer absorption band of the dye. The dye–CB6 assembly has the potential for the detection of biogenic amines under physiological pH conditions and at low analyte concentrations.

Chlorination of tramadol: Reaction kinetics, mechanism and genotoxicity evaluation

The major foodborne pathogens have caused significant foodborne illness and even death, while spoilage microorganisms lead to remarkable public health issues worldwide. To address these concerns, combination of the natural preservative with potential organic acid is the superior solution because the use of synthetic preservatives alone is aware of health impacts to consumers. Therefore, this research aimed to identify the combined effect of these two substances against pathogenic and spoilage bacterial strains in different pH conditions (non-adjusted, mildly acidic, and neutral pH conditions). Eighteen ethanol plant extracts with five organic acids were used in this study to test for antibacterial activities through Disk Diffusion method to determine the zone of inhibition, and plate dilution with Broth Dilution method to determine the MIC and MBC respectively. The potential plant extract and organic acid that demonstrated stronger activity were selected for the combinatorial study. In all tested pH conditions, clove extract demonstrated high potency when lactic acid or acetic acid was incorporated. At mildly acidic conditions, combinations were more effective in reducing the MICs of clove extract combined with acetic acid against all tested strains. The combination resulted as a synergistic effect with the FIC index of 0.5 for the test on Listeria monocytogenes 4b. Therefore, the amplified activity was established when clove extract combined with acetic acid. The obtained data was evaluated to have a combinatorial effect, which could be a potential object for controlling pathogenic and spoilage bacteria.