Sustainable and practical degradation of intact chicken feathers by cultivating a newly isolated thermophilic <Emphasis Type="Italic">Meiothermus ruber</Emphasis> H328

The sustainable and practical degradation of intact chicken feathers by a newly isolated thermophilic bacterium Meiothermus ruber H328 was presented with extensive data. Aerobic cultivation with moderately thermophilic strain H328 at 55°C for 6 days led to the apparently complete decay of the truly intact feathers and provided 1.89 mmol free amino acids and 7.32 mmol acid-hydrolyzed amino acids from 50 ml of culture containing 3% (w/v) intact chicken feathers. The amino acid components in the soluble fraction of the culture conspicuously agreed with those calculated from the intact feathers. This demonstrated that more than 55% of total keratin proteins were solubilized from the intact chicken feathers into the culture in the forms of free amino acid and/or soluble oligopeptide, and most of them are directly derived from the intact feathers by proteolytic digestion. Feather degradation by strain H328 surpasses that by any other microorganisms with regard to degradation efficiency, absence of requirement for pretreatment of the feathers, and product fidelity in the amino acid component. Furthermore, the culture containing the degradative products from the intact feathers was subjected to matrix-assisted laser desorption ionization mass spectrometry-time-of-flight analysis, and it was revealed that the molecular masses of the solubilized products, oligopeptides, were less than 1,000. This result allows us to investigate the bioactivities of oligopeptides derived from the degradation of chicken feathers by cultivation with strain H328 as well as the production of amino acids for feedstuffs.     

Synthesis of Nucleoside <Emphasis Type="Italic">N</Emphasis>-Phosphoamino Acids and Peptide Formation

Nucleoside N-phosphoamino acids were synthesized through Atherton-Todd reaction of nucleoside H-phosphonate with amino acids, and their structures were confirmed by NMR and ESI-MS. After nucleoside N-phosphoamino acid was incubated in anhydrous methanol at 40 °C for 72 h, di- to tetra-peptide derivatives were detected by ESI-MS, and their structures were further identified by multistage mass spectrometry. These and previously published studies in aqueous solution suggest that nucleoside N-phosphoamino acids could have been prebiotic precursors of oligopeptides.     

Oligopeptide assimilation and transport by Oenococcus oeni

Aims: Oenococcus oeni is a slow‐growing wine bacterium with a low growth yield. It thrives better on complex nitrogen sources than on free amino‐acid medium. We aimed to characterize the oligopeptide use of this micro‐organism. Methods and Results: Several peptides of two to eight amino‐acid residues were able to provide essential amino acids. The disappearance of various peptides from extracellular medium was assessed with whole cells. Initial rates of utilization varied with the peptide, and free amino acids were released into the medium. Conclusions: Oenococcus oeni was able to transport the oligopeptides with two to five amino‐acid residues tested and to hydrolyse them further. Significance and Impact of the Study: This study has clear implications for the relationship between wine nitrogen composition and the ability of O. oeni to cope with its environment.     

Peptide transport by germinating barley embryos: Uptake of physiological di- and oligopeptides

The uptake of a variety of physiological di- and oligopeptides by germinating barley (Hordeum vulgare L.) embryos is described. Peptides as large as pentaalanine can be absorbed. Evidence is presented suggesting the peptides are absorbed intact and subsequently undergo rapid intracellular hydrolysis. Uptake shows stereospecificity. The transport of peptides is generally faster than the transport of amino acids, making it likely that the former could play an important role in the mobilization of the protein storage reserves during germination. The peptide transport system in barley is compared with similar systems from other groups of organisms.Abbreviations Gly-sar glycylsarcosine - Gly-sar-sar glycylsarcosylsarcosine - Gly-sar-sar-sar Glycylsarcosylsarcosylsarcosine     

Oenococcus oeni preference for peptides: qualitative and quantitative analysis of nitrogen assimilation

Optimization of malolactic fermentation in wine depends mainly on better understanding of nitrogen nutritional requirements of Oenococcus oeni. Four widely used starter strains and the reference ATCC BAA-1163 strain were grown in media containing different N sources: free amino acids, oligopeptides (0.5–10 kDa) or polypeptides (> 10 kDa). Amino acid auxotrophies were determined by the single omission technique. The tested strains were indifferent to only two to four amino acids and two of the starter strains appeared to be particularly demanding. Nitrogen consumption was investigated and a significant level of nitrogen was consumed by O. oeni only in the free amino acid medium. In media containing complex nitrogen sources, a global balance above 5 mg N l⁻¹ was enough to ensure biomass formation of all tested strains. Moreover, for all strains, bacterial growth yield was higher in the presence of nitrogen from peptides than that from free amino acids. However, no direct relationship between the bacterial growth level and the amount of nitrogen metabolized could be established. These findings were discussed in relation to the physiology of wine malolactic bacteria.     

Components Contributing to the Improvement of Meat Taste during Storage

The changes in both taste and taste components of beef, pork, and chicken during storage were examined.

The brothy taste intensity of pork and chicken was significantly stronger after conditioning than before. On the other hand, for beef, there was no significant difference in the brothy taste intensity before or after conditioning. The analysis of major taste components showed that the levels of free amino acids in all meats were higher after conditioning than before. The differences in the levels of free amino acids before versus after conditioning were large in pork and chicken and very small in beef. Oligopeptide levels were lower in beef after conditioning than before, but they were higher in pork and chicken after conditioning than before. These results corresponded to results of the sensory evaluation studies described above, indicating that free amino acids and oligopeptides contributed to the improvement of meat taste during storage.     

Peptide transport by germinating barley embryos: Evidence for a single common carrier for di- and oligopeptides

Competition for uptake of a range of amino acids and peptides by germinating barley (Hordeum vulgare L.) embryos was studied. Peptides and amino acids show no competition and are apparently absorbed by independent transport systems. However, peptides of widely different structures do compete and it seems that only a single peptide transport system is present in barley embryos, capable of handling both di- and oligopeptides. The ability of physiological peptides to totally inhibit the uptake of glycylsarcosine indicates they share a common uptake system which previously has been shown to have the properties of an active transport process. The characteristics of the barley peptide transport system are compared with those found in other organisms.     

Peptide synthesis using o-nitrophenylsulfenyl N-carboxy alpha-amino acid anhydrides. Sequential oligopeptides having alternate gamma-methyl L-glutamyl and L-phenylalanyl residues.

A series of sequential oligopeptides having the sequence alternating γ-methyl L -glutamyl and L -phenylalanyl residues have been successfully prepared by a rapid method involving the reaction of o-nitrophenylsulfenyl N-carboxy α-amino acid anhydrides with amino acid and peptide esters. The sequential oligopeptides, which are interesting from a conformational aspect, were obtained in optical pure forms above 74% yields. This result demonstrates that the o-nitrophenylsulfenyl N-carboxy α-amino acid anhydride method is especially useful for easy synthesis of protected oligopeptides with o-nitrophenylsulfenyl group.     

Food protein fragments are regulatory oligopeptides

Until recently food proteins were considered to be an energy source and a source of essential and nonessential amino acids required for protein synthesis and precursors of many vital biomolecules. However, we assumed earlier that food protein fragments might perform some regulatory functions. The theoretical justification for this assumption is advanced in this work. In the present work, the primary structures of protein fragments were compared with amino acid sequences of known natural regulatory oligopeptides in silico. It is shown that fragments formed as a result of animal food protein cleavage by proteolytic enzymes can exist in the gastrointestinal tract for a long time. Many of them are enzyme inhibitors, regulators of nervous, endocrine, and immune system, and possess antimicrobial and other activities. It has also been shown that the lifetime of fragments before their cleavage in the gastrointestinal tract could be enough for performing corrective functions. Thus, as a result of food protein fragmentation a dynamic pool of exogenous regulatory oligopeptides with functions changing as shorter fragments are generated may form. The detection of an endogenous-exogenous pool of regulatory molecules expands the significance and content of the Ashmarin-Obukhova hypothesis on a functional continuum of natural oligopeptides. The possible practical importance of these results is noted.     

Role of Pre-epithelial ``Unstirred' Layers in Absorption of Nutrients from the Human Jejunum

Pre-epithelial ``unstirred' layers (abbr. pL) are generally regarded as undesirable diffusion barriers that impede access to absorptive cells of exogenous hexoses, amino acids or other experimental probes added to fluids bathing the mucosal surface. In the present paper it is suggested that the pL may have a functional role. Diffusion plus convection of saccharides and oligopeptides from lumen to brush border, combined with absorption to blood of their hydrolytic products, confers rectifying properties to the pL. The proposed model, based on experimental data from segmental jejunal perfusions in normal human subjects, indicates that the functional pathway for diffusion plus convection through the pL of hexoses and amino acids bound in the form of oligomers is only 10 ± 2 μm or little more than the anatomical thickness of the glycocalyx and mucus layers. In contrast, the pathlength from brush border to lumenal perfusion fluid for diffusion minus convection of monomers generated by membrane bound hydrolases is 50–150 μm. According to this model the pL offers little resistance to the passage of saccharides or oligopeptides from lumen to brush border but at the same time it provides a protective blanket that diminishes diffusional losses to lumenal chyme of hexoses and amino acids generated in the brush border. The model provides a theoretical explanation for the ``kinetic advantage' of transporting hexoses or amino acids through the pL in the form of oligomers and it predicts the proximal-distal concentrations of free glucose or fructose found experimentally in the outflows from jejunal segments perfused with sucrose or maltose. Received: 5 September 2000/Revised: 31 October 2000     

Free amino acids in Phycomyces blakesleeanus during development

In Phycomyces blakesleeanus the total level of free amino acids and their relative distributions change strikingly during the development of sporangiophores. Very high levels of serine, glutamate and histidine are found. The variety of amino acids becomes more restricted in sporangiophores grown on nitrogen-rich media; levels of arginine increase dramatically. The distribution differs in mycelia. No unusual amino acids or oligopeptides were found.     

Convenient solid-phase synthesis of oligopeptides using pentacoordinated phosphoranes with amino acid residue as building blocks

The reactive intermediates of pentacoordinated phosphoranes with amino acids (P(5)-AA) as building blocks, which were obtained by the reaction of O-phenylene phosphorochloridate with N,O-bis(trimethylsilyl)amino acids, were linked to a solid-phase support containing a hydroxymethyl polystyrene functional group. The first amino acid residue was coupled to the solid-phase support after washing the resin with organic solvent. Repeating the procedure led to oligopeptides linked on the resin. A series of free oligopeptides including tetra-Gly, di-Val, tri-Val, di-Leu, di-Phe, and Phe-Leu were obtained after cleavage from solid-phase support. The structure of these oligopeptides were determined by IR, (1)H NMR, FAB-MS, and HPLC.     

MUCOPOLYSACCHARIDES PRODUCED BY A STRAIN OF CLOSTRIDIUM PERFRINGENS.

Izumi, Kunihiko (Kanazawa University, Kanazawa, Japan). Mucopolysaccharides produced by a strain of Clostridium perfringens. J. Bacteriol. 83:956-959. 1962.-A new series of mucopolysaccharides was isolated from the culture medium of Clostridium perfringens and partially purified by the use of a column of anion-exchange resin. A large part of the substance was composed of neutral sugars, amino sugars, uronic acids, and oligopeptides, suggesting a structure analogous to that of bacterial cell walls. Acidic amino acids, especially aspartic acid, were the main constituents of the oligopeptides. The substance exhibited high viscosity when dissolved in water. The degree of viscosity in each fraction seemed to depend on the content of amino sugars and the chain length of the oligopeptides.     

Chemotaxis of the unicellular green algaDunaliella salina and the ciliatedTetrahymena pyriformis—effects of glycine,lysine, and alanine,and their oligopeptides

Chemotactic properties of amino acids (L-alanine, glycine and L-lysine) and their oligopeptides (10^–6M) and binding sites to these ligands were investigated in two unicellular models, the heterotrophicTetrahymena pyriformis and the auxotrophicDunaliella salina. Chemotaxis ofDunaliella induced by simple amino acids and their derivatives demonstrated that binding sites (receptors) for food molecules are not only present in the membrane but are also able to induce their basic physiological response. InTetrahymena, substances with special molecular structure and properties (polar, hydrophilic character of the signal peptide chain)-5-L-Lys, 5-Glywere required for chemoattraction, other peptides tested, lacking the required structure, were repellent. Divergences in chemotaxis and binding assays of both species suggest that trends of functional and binding parameters do not run parallel at this level of evolution.     

Self-organization of oligopeptides obtained on dissolution of feather keratins in superheated water

Keratins are self-organized proteins that are abundantly available in wool, feather, human hair, etc., making them a potential cheap feedstock for the modification of amino acids. This paper explores the hydrolysis of keratin in water under specific pressure-temperature conditions where the hydrolysis through scission of the protein chain yields oligopeptides. Here we report for the first time that, under appropriate conditions, these oligopeptides self-assemble into a hierarchical architecture, the process being followed in time by optical microscopy. Birefringent needle-like crystals are observed which tend to nucleate heterogeneously. When given sufficient time, these needles become tens of microns in length and act as further nuclei, developing a highly repetitive structure of several hundreds of microns in size. Micro-focus X-ray diffraction studies supported by in situ microscopy reveal that these needles have a crystal structure similar to that of the native protein, although better organized along the ab-plane. Spectroscopic studies on these structures show crystalline bands that disappear above 150 degrees C, coinciding with an endothermic peak in DSC. Amino acid analysis shows that the self-assembled birefringent entities are indeed oligopeptides, consisting of sequences of approximately 40 amino acids. The proposed ecofriendly route provides an effective route for obtaining oligopeptides that can be used as important building blocks for the synthesis of a range of novel polymers. The oligopeptides obtained from the sustainable source can be used as important building blocks for the synthesis of a range of novel polymers.     

Conformation of oligopeptides with L-leucyl-L-leucyl-L-alanyl and L-methionyl-L-methionyl-L-leucyl repeating units

The conformation of oligopeptides with hydrophobic side chains, Nps-(L -Leu-L -Leu-L -Ala)_n-OEt and Nps-(L -Met-L -Met-L -Leu)_n-OEt(n = 1–6), in the solid state, obtained either by evaporation of the solvent or by precipitation with diethyl ether from a 1,1,1,3,3,3-hexafluoropropan-2-ol (HFIP) solution, has been studied with ir spectroscopy and x-ray powder-diffraction measurements. The conformation of these peptides in the HFIP solution has been studied by CD spectroscopy. Due to a strong preference of the amino acids to form an α helix, the peptides begin forming α helices at the dodecapeptide in the HFIP solution, and in the solid state by evaporation. In the solid state, with precipitation, the α-helical conformation is first observed at the octadecapeptide and the lower peptides assume a β structure. The conformational change, from the α helix to the β structure of the peptides with 12 to 15 amino acid residues, during the precipitation process, is due to a strong tendency of the amino acids to form the β-structure in rather short peptide lengths.     

A novel and efficient enzymatic method for the production of peptides from unprotected starting materials

We report herein the development of a novel and efficient enzymatic method for the production of oligopeptides. This newly discovered method is a simple, cost-effective process, using unprotected amino acids as substrates in an aqueous solution and producing peptides in high yield. The target of our initial screen was l-alanyl-L-glutamine, a dipeptide of significant industrial interest by virtue of its widespread use in infusion therapy. By means of the screening of microorganisms that can catalyze the peptide-forming reaction producing l-alanyl-L-glutamine from L-alanine methylester (acyl donor) and L-glutamine (nucleophile), we discovered that Empedobacter brevis ATCC 14234 produced l-alanyl-L-glutamine most efficiently. The newly found enzyme purified from E. brevis ATCC 14234 facilitates significantly high production yields of l-alanyl-L-glutamine from L-alanine methylester and L-glutamine in an aqueous solution--more than 80% yield based on L-alanine methylester. In addition, this enzyme has wide substrate specificity--both for acyl donors and nucleophiles--and can catalyze peptide-forming reactions not only to produce various dipeptides from the corresponding amino acid esters and amino acids but also to produce various oligopeptides from the corresponding amino acid esters and peptides.     

Oxyma‐based phosphates for racemization‐free peptide segment couplings

Glyceroacetonide–Oxyma [(2,2‐dimethyl‐1,3‐dioxolan‐4‐yl)methyl 2‐cyano‐2‐(hydroxyimino)acetate ( 1 )] displayed remarkable physico‐chemical properties as an additive for peptide‐forming reactions. Although racemization‐free amide‐forming reactions have been established for N‐urethane‐protected α‐amino acids with EDCI, 1 , and NaHCO₃ in water or DMF‐water media, amide‐forming reactions of N‐acyl‐protected α‐amino acids and segment couplings of oligopeptides still require further development. Diethylphosphoryl–glyceroacetonide–oxyma (DPGOx 3 ) exhibits relative stability in aprotic solvents and is an effective coupling reagent for N‐acyl‐protected α‐amino acids and oligo peptide segments. The conditions reported here is also effective in lactam‐forming reactions. Unlike most of the reported coupling reagents, simple aqueous work‐up procedures can remove the reagents and by‐products generated in the reactions. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

Analysis of cytoplasmic membrane proteome of Streptococcus pneumoniae by shotgun proteomic approach

In this study, cytoplasmic membrane proteins of S. pneumoniae strain R6 (ATCC BBA-255) were effectively separated from cell wall or extracellular proteins by sodium carbonate precipitation (SCP) and ultracentrifugation. Forty seven proteins were analyzed as cytoplasmic membrane proteins from the 260 proteins identified by the shotgun proteomic method using SDS-PAGE/LC/MS-MS. ABC transporters for metabolites such as metals, oligopeptides, phosphate, sugar, and amino acids, and membrane proteins involved in phosphotransferse systems, were identified as the predominant and abundant, cytoplasmic membrane proteins that would be essential for nutrient uptake, antibiotic resistance and virulence mechanisms. Our result supports that gel-based shotgun proteomics combined with sodium carbonate precipitation and ultracentrifugation is an effective method for analysis of cytoplasmic membrane proteins of S. pneumoniae.     

Oligopeptides as plant growth regulators

It is generally accepted that plant growth and development are regulated by the known plant hormones. Some objections to the functions of auxins and cytokinins in the induction of shoot and root primordia are reported. Instead of them oligopeptides of special amino acid sequences could be the endogenous signals. There exist structure relationships between auxins and parts of the α-helical oligopeptides of defined amino acid sequences. The same is true for cytokinins. The most difficult part of this hypothesis is its verification. Using protonemata ofFunaria hygrometrica bud induction by various oligopeptides was investigated. The most active peptide tested is leucine-tryptophan. On the other hand endogenous oligopeptides containing [¹⁴C]-leucine in the moss protonemata during endogenous bud initiation were looked for. Three to four different oligopeptide spots seem to be related to bud induction.