What makes a good graphene-binding peptide? Adsorption of amino acids and peptides at aqueous graphene interfaces

Investigation of the non-covalent interaction of biomolecules with aqueous graphene interfaces is a rapidly expanding area. However, reliable exploitation of these interfaces in many applications requires that the links between the sequence and binding of the adsorbed peptide structures be clearly established. Molecular dynamics (MD) simulations can play a key role in elucidating the conformational ensemble of peptides adsorbed at graphene interfaces, helping to elucidate these rules in partnership with experimental characterisation. We apply our recently-developed polarisable force-field for biomolecule-graphene interfaces, GRAPPA, in partnership with advanced simulation approaches, to probe the adsorption behaviour of peptides at aqueous graphene. First we determine the free energy of adsorption of all twenty naturally occurring amino acids (AAs) via metadynamics simulations, providing a benchmark for interpreting peptide-graphene adsorption studies. From these free energies, we find that strong-binding amino acids have flat and/or compact side chain groups, and we relate this behaviour to the interfacial solvent structuring. Second, we apply replica exchange with solute tempering simulations to efficiently and widely sample the conformational ensemble of two experimentally-characterised peptide sequences, P1 and its alanine mutant P1A3, in solution and adsorbed on graphene. For P1 we find a significant minority of the conformational ensemble possesses a helical structure, both in solution and when adsorbed, while P1A3 features mostly extended, random-coil conformations. In solution this helical P1 configuration is stabilised through favourable intra-peptide interactions, while the adsorbed structure is stabilised via interaction of four strongly-binding residues, identified from our metadynamics simulations, with the aqueous graphene interface. Our findings rationalise the performance of the P1 sequence as a known graphene binder.

Relationships of nicotianamine and other amino acids with nickel, zinc and iron in Thlaspi hyperaccumulators

Experimental evidence suggests that nicotianamine (NA) is involved in the complexation of metal ions in some metal-hyperaccumulating plants. Closely-related nickel (Ni)- and zinc (Zn)-hyperaccumulating species were studied to determine whether a correlation exists between the Ni and Zn concentrations and NA in foliar tissues. A liquid chromatography–mass spectrometry (LC-MS) procedure was developed to quantify the NA and amino acid contents using the derivatizing agent 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate. A strong correlation emerged between Ni and NA, but not between Zn and NA. Concentrations of NA and l-histidine (His) also increased in response to higher Ni concentrations in the hydroponic solution supplied to a serpentine population of Thlaspi caerulescens. An inversely proportional correlation was found between the iron (Fe) and Ni concentrations in the leaves. Correlations were also found between Zn and asparagine. The results obtained in this study suggest that NA is involved in hyperaccumulation of Ni but not Zn. The inverse proportionality between the Ni and Fe concentrations in the leaf may suggest that Ni and Fe compete for complexation to NA.

Simultaneous analysis of free amino acids and biogenic amines in honey and wine samples using in loop orthophthalaldeyde derivatization procedure

This work presents a RP-HPLC method for the simultaneous quantification of free amino acids and biogenic amines in liquid food matrices and the results of the application to honey and wine samples obtained from different production processes and geographic origins. The developed methodology is based on a pre-column derivatization with o-phthaldialdehyde carried out in the sample injection loop. The compounds were separated in a Nova-Pack RP-C18 column (150 mm × 3.9 mm, 4 μm) at 35 °C. The mobile phase used was a mixture of phase A: 10 mM sodium phosphate buffer (pH 7.3), methanol and tetrahydrofuran (91:8:1); and phase B: methanol and phosphate buffer (80:20), with a flow rate of 1.0 ml/min. Fluorescence detection was used at an excitation wavelength of 335 nm and an emission wavelength of 440 nm. The separation and quantification of 19 amino acids and 6 amines was carried out in a single run as their OPA/MCE derivatives elute within 80 min, ensuring a reproducible quantification. The method showed to be adequate for the purpose, with an average RSD of 2% for the different amino acids; detection limits varying between 0.71 mg/l (Asn) and 8.26 mg/l (Lys) and recovery rates between 63.0% (Cad) and 98.0% (Asp). The amino acids present at the highest concentration in honey and wine samples were phenylalanine and arginine, respectively. Only residual levels of biogenic amines were detected in the analysed samples.

Sunflower meal in commercial layer diets formulated on total and digestible amino acids basis

An experiment was conduced to evaluate the inclusion of sunflower meal (SBM) in commercial layer diets formulated on total or digestible amino acids basis. One hundred forty-four 41-week-old Lohmann LSL layers were distributed in a completely randomized experimental design in a 2 x 4 factorial arrangement with three replications of six birds each. Treatments consisted of a combination of four SBM inclusion levels SBM(0%, 4%, 8%, and 12%) and feed formulation according two amino acid recommendations (total or digestible). The experimental period was divided into five periods of fourteen days. Performance parameters (egg production, feed intake, feed conversion, egg mass) were evaluated for each period. In the last two days of each period, three eggs per replication were collected to evaluate egg quality (Haugh units, specific gravity, egg weight, eggshell thickness, and eggshell percentage). Hens fed on total amino acid recommendation presented the highest values for egg weight. Diets formulated on digestible amino acids basis showed an improvement in eggshell percentage and egg specific gravity. SBM addition in commercial layer diets did not influence performance; however, increasing SBM dietary levels SBM improved eggshell quality.

Commercial laying hen diets formulated according to different recommendations of total and digestible amino acids

An experiment was conducted to evaluate different commercial laying hen diets formulated based on recommendations for total and digestible amino acids. One hundred and twenty Lohmann LSL commercial laying hens aged 25 weeks were distributed in a completely randomized experimental design involving five replications of six birds in four treatments. Diet formulation on a total amino acid basis followed the recommendations of NRC (1994) and Rostagno et al. (2000), whereas formulation on digestible amino acids basis was according to Rostagno et al. (2000) and Degussa (1997) recommendations. The experimental period was divided into five periods of fourteen days. Performance parameters (egg production, feed intake, feed conversion, egg mass) were evaluated for each period, and on the last two days of each period, three eggs per replication were collected to evaluate egg quality parameters (Haugh unit, egg specific gravity, egg weight, eggshell thickness and percentage). Means were compared by orthogonal contrasts. Results on feed intake, egg production, egg mass, feed conversion and egg specific gravity showed that total amino acid recommendations promoted better bird responses than digestible amino acid recommendations.

Synthesis of potentially bioactive PABA-related N-(aminoalkyl)lactamic amino acids and esters via selective SNAr reactions

Potentially bioactive N-(aminoalkyl)lactamic amino acids and esters were synthesized in satisfactory to good yields by SNAr reactions of aromatic acids with N-(3-aminopropyl)lactams followed by esterification with tertiary amino alcohols. The addition-elimination SNAr mechanism was confirmed by NMR and MS measurements.

Mechanisms of insulin secretion in malnutrition: modulation by amino acids in rodent models

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Photoacoustic spectroscopy of aromatic amino acids in proteins

This paper concerns the use of photoacoustic spectroscopy (PAS) to study the presence of aromatic amino acid in proteins. We examined the aromatic amino acids in six proteins with well-known structures using absorption spectra of near ultraviolet PAS over the wavelength range 240-320 nm. The fundamental understanding of the physical and chemical properties that govern the absorption of light and a subsequent release of heat to generate a transient pressure wave was used to test the concept of monitoring aromatic amino acids with this method. Second derivative spectroscopy in the ultraviolet region of proteins was also used to study the regions surrounding the aromatics and the percentage area in each band was related in order to determine the contribution in function of the respective molar extinction coefficients for each residue. Further investigation was conducted into the interaction between sodium dodecyl sulphate (SDS) and bothropstoxin-I (BthTx-I), with the purpose of identifying the aromatics that participate in the interaction. The clear changes in the second derivative and curve-fitting procedures suggest that initial SDS binding to the tryptophan located in the dimer interface and above 10 SDS an increased intensity between 260 and 320 nm, demonstrating that the more widespread tyrosine and phenylalanine residues contribute to the SDS/BthTx-I interactions. These results demonstrate the potential of near UV-PAS for the investigation of membrane proteins/detergent complexes in which light scattering is significant.

The activity of amino acids produced by Paenibacillus macerans and from commercial sources against the root-knot nematode Meloidogyne exigua

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Electrochemical behavior of the N-nosyl-protected amino acids in N,N-dimethylformamide

The electrochemical reduction of serine, glycine, and leucine protected by the 4-nitrobenzenesulfonyl, group in N,N-dimethylformamide at mercury cathode occurs at two steps. The first one at -0.8 V vs. SCE, after a one-electron transfer, leads the anion radical formation that dimerizes and adsorbs at electrode. In the second step at -1.4 V, an instable dianion forms which then cleaves. The mechanism is discussed.

Molecular orbital calculation for the model compounds of kainoid amino acids, agonists of excitatory amino acid receptors. Does the kainoid C4-substituent directly interact with the receptors?

Kainoid amino acids are agonists of the AMPA/kainate receptors and exhibit highly potent neuroexcitatory activity. From the results of extensive structure-activity relationship studies, we previously postulated that the C4-substituent of the kainoid amino acids interacts with an allosteric site of the glutamate receptor with electron-donating character. In order to investigate the mode of action in more detail, molecular orbital calculation for model compounds of the kainoid were performed. The results indicated that the HOMO energy level of the C4-substituent is involved in the potent neuroexcitatory activity, thus supporting our hypothesis. (C) 2002 Elsevier B.V. Ltd. All rights reserved.

Influence of free amino acids on clavulanic acid production by Streptomyces clavuligerus in synthetic medium

The effect of nitrogen source on clavulanic acid production was investigated in shake flasks. Media containing asparagine plus one of several different amino acids or a combination of two amino acids was tested. The best result, ca. 180 mg/L clavulanic acid, CA, in 60 h, was obtained with the lysine-tyrosine pair In an aerated and agitated fermentor this medium led to CA concentrations of ca. 210 mg/L, a remarkable production for synthetic medium utilization. Amino acids analysis during cultivation indicated that, while asparagine was consumed rapidly, lysine and tyrosine were metabolized slowly, promoting CA production.

Association of free amino acids with caries experience and mutans streptococci levels in whole saliva of children with early childhood caries

Objective: The aim of the present study was to identify the free amino acid content in whole saliva of children with (CE) and without early childhood caries (CF) (ECC), correlating these findings with caries experience and mutans streptococci (MS) levels in saliva.Design: Seventy-eight healthy children, both genders, 6-71 months age, were selected to participate in the study. Following examination for dmft scores calculation, unstimulated whole saliva was collected from all 78 participants, stored at -80 degrees C, and used for amino acid analysis, on a Biochem 20 plus amino acid analyzer. Stimulated whole saliva was collected from 52 children, transported, diluted and plated on MSB agar medium for detection of MS in cfu/mL.Results: Forty different free amino acids were identified in whole saliva, with great variation in their concentration. A statistically significant relation was found between caries experience and the presence of free proline and glycine. While proline (p = 0.0182) was more frequently absent in the CF group, the absence of glycine (p = 0.0397) was more often observed in the CE group. In the presence of higher levels of MS, free glycine reduced the risk of experiencing dental caries (p = 0.0419). Conversely, the presence of proline was found to increase the risk of experiencing the disease (p = 0.0492).Conclusions: The presence of free proline and absence of free glycine in children with ECC, highly contaminated with MS, increased the chances of experiencing dental caries in the present population. Further studies are needed to better understand this phenomenon. (C) 2008 Elsevier Ltd. All rights reserved.

Apparent digestibility of protein, energy, and amino acids in some selected feed ingredients for pacu Piaractus mesopotamicus

The apparent digestibility coefficients (ADCs) of amino acids (AA), protein, and energy in six feed ingredients (Brazilian fish meal, soybean meal, corn gluten meal, alcoholic yeast, corn, and wheat bran) we evaluated for pacu juveniles. In general, all ingredients showed high digestibility values for all AA, and differences among ADCs of individual amino acid were detected (P < 0.01). Corn gluten, soybean, and fish meals had the highest ADCs of AA. The ADCs of protein in fish meal, yeast, and corn gluten meal should not be used as AA digestibility indicators, because those showed differences up to 6.7% between the ADCs of protein and AA. All ingredients had lower ADCs of energy than corn gluten meal (P < 0.01). Lysine was the first limiting amino acid in fish meal, corn gluten meal, wheat bran, and corn, and the second limiting amino acid in soybean meal, as methionine was the first limiting amino acid in soybean meal and yeast. However, the soybean meal was the best quality protein source, as it had the highest digestible essential amino acid index. This demonstrated that digestible amino acid values can be used to formulate practical diets for pacu, preventing potential deficiencies or excess that might cause environmental and economic losses.