Critical chain length for helix formation in L-methionine oligopeptides

The circular dichroism of a series of L -methionine oligopeptides [BOC-(Met)_n-OMe] was examined in trifluoroethanol and hexafluoroacetone sesquihydrate. The results indicate that the trimer through the hexamer exists predominantly in disordered conformations in these solvents. An abrupt change in the CD pattern at the heptamer in trifluoroethanol suggests that L -methionine oligopeptides begin forming helices at this chain length.     

Sequential oligopeptides. Synthesis and characterization of the oligopeptides and a polypeptide with the repeating sequence L-norvalyl-glycyl-L-proline

The synthesis and characterization of a series of oligopeptides (from the tripeptide to the octadecapeptide) with the repeating sequence L -norvalyl-glycyl-L -proline and a polytripeptide with this sequence are reported. The oligomers were synthesized step by step using the mixed anhydride method. All the products were chemically and optically pure. The polymer was prepared by the active ester method, using the p-nitrophenyl ester as the polymerizable tripeptide derivative. Good yield of relatively high average molecular-weight polymer was obtained. In the accompanying paper conformational investigations, both in solution and in the solid state, on the oligomers and the polymer are described.     

Circular dichroism studies of helical oligopeptides. Can 3(10) and alpha-helical conformations be chiroptically distinguished?

Circular dichroism studies of seven helical oligopeptides containing alpha-aminoisobutyric acid (Aib) in methanol and trifluoroethanol (TFE) solutions are reported. Peptides ranging from 10 to 21 residues in length have been examined. In all cases distinct negative CD bands characteristic of helical peptides are obtained at approximately 220 and 205 nm corresponding to the n-pi and pi-pi transitions, respectively. The ratio R = [theta] pi-pi is less than 1.0 for all peptides studied. Using crystal structure and n.m.r. results for a 10 residue 3(10) helical peptide and literature values for an alpha-helical 11-residue peptide, it is shown that both helical conformations yield R values of approximately 0.8 in alcoholic solvents. The CD data are considered in the light of 1H n.m.r. studies on these oligopeptides. The results suggest that 3(10) and alpha-helical conformations cannot be distinguished by CD methods.     

Circular dichroism studies of isoleucine oligopeptides in solution

A conformational analysis was carried out on a series of L -isoleucine oligomers having the general formula BOC-(Ileu)_n-OMe (n = 2–8). These oligopeptides were examined in trifluoroethanol, trifluoroethanol-acid and mixed organic-water media. The results indicate that these oligomers form β-conformations beginning at the heptamer. The stability of the β-conformations was found to be greater than those formed by oligopeptides derived from L -alanine.     

Fragmentomics of proteins and natural oligopeptides

The term fragmentomics is grounded and defined. Theoretical structure-function analysis of all possible fragments of a protein molecule was performed under the concept of fragmentomics to determine the regions that could be potential sources of regulatory oligopeptides. For this purpose, we used the data on the primary structure of bovine hemoglobin, the information contained in the EROP-Moscow database on the structures and functions of natural oligopeptides, and a specialized software package. This analysis revealed natural nonhemoglobin oligopeptides containing hemoglobin fragments and natural oligopeptides with the structure precisely coinciding with hemoglobin fragments. The most abundant of them are neuropeptides, antimicrobial oligopeptides, and hormones. It was demonstrated that the tetrapeptide and larger fragments of hemoglobin identified in nonhemoglobin oligopeptides and possessing a mentioned activity are present in the amino acid sequences of experimentally determined hemoglobin oligopeptides with the same function. The proposed approach allowed us to discover new potentially active sites in the hemoglobin amino acid sequence not yet studied experimentally. The possibility of natural formation of regulatory oligopeptides from hemoglobin molecules and other food proteins is discussed, as well as the generation of an exogenous oligopeptide pool in the gastrointestinal tract, and how the results match the concept of natural continuum of regulatory oligopeptides.     

Conformational aspects of polypeptides. XVIII. Conformational studies of oligopeptides derived from L-alanine

The conformations of oligopeptides derived from L -alanine and co-oligomers of L -alanine with γ-methyl-L -glutamate were studied in several solvents via optical rotation and far-ultraviolet spectroscopy. Calculated values for optical rotation based on model compounds were compared with experimental values for the oligomers. In trifluoroacetic and dichloroacetic acids, the oligomers and co-oligomers exhibit rotations in close agreement with predicted values based on model compounds. Thus, in these solvents only nonhelical conformations exist. In trifluoroethanol, the experimental points of molar rotation for the pentamer and larger oligomers no longer follow the predicted values. In addition, the benzyloxycarbonyl and acetyl cononamers show b₀ values of about ?150, which demonstrates the presence of stable helical forms for these peptides. We also examined the molar extinction coefficients of oligopeptides in the 190 mμ region and determined the values for nonhelical peptide groups. The molar extinction coefficients per amide bond for the benzyloxycarbonyl and acetyl cononamers show extensive hypo-chromism, once again indicating the presence of stable helices for these compounds in trifluoroethanol.     

Synthesis and conformational study in solution and in solid state of oligopeptides containing L-leucine and glycine

The synthesis and conformational studies of the oligopeptide N-tert.-butyloxycarbonyl-L-Leu-(L-Leu-Gly)n-OBzl (n = 1, 3, 5) and N-tert.-butyloxycarbonyl-(L-Leu-Gly)2-OBzl are described. The peptides were synthesized by stepwise and fragment condensation techniques using dicyclohexylcarbodiimide as the condensing agent in solution. The conformational study of the oligopeptides was carried out using CD, u.v. and i.r. spectra. The conformation in solution was examined in trifluoroethanol, hexafluoroisopropanol, hexafluoroacetone trihydrate, and methanol. CD spectra in trifluoroethanol exhibited a gradual variation with increasing peptide chain length. This can be interpreted as a formation of an ordered structure which is already present in the heptapeptide and, to a greater extent, in the undecapeptide. The results obtained from the CD profiles and i.r. spectra showed the presence of beta structure with antiparallel chains in the heptapeptide and undecapeptide. Finally, CD spectra revealed in trifluoroethanol-water solution the binding of Ca2+ to heptapeptide and undecapeptide together with a contemporaneous conformational change. This change is probably due to the formation of beta-turns. No change in the CD profiles was obtained by using Mg2+, K+, Na+, and Li+ ions instead of Ca2+.     

Immunological properties of uricase conjugated to neutral soluble polymers.

For a comparative study of immunological properties of protein-polymer conjugates, uricase was modified with (a) poly(N-vinylpyrrolidone) 6000 Da, (b) poly(N-acriloylmorpholine) 6000 Da, (c) branched monomethoxypoly(ethylene glycol) 10000 Da, and (d) linear monomethoxypoly(ethylene glycol) 5000 Da. Spectroscopic studies performed by UV, fluorescence, and circular dichroism did not show any relevant difference in protein conformation among the native and the conjugates. Immunological studies showed that both uricase antigenicity and immunogenicity were altered by polymer conjugation to an extent that depended upon the polymer composition; in particular, monomethoxypoly(ethylene glycol) 10000 Da remarkably reduced the protein antigenicity, while unexpectedly, the poly(N-vinylpyrrolidone) derivative presented higher antigenicity than the native protein. In Balb/c mice, the native protein elicited a rapid and intense immunoresponse whereas all the conjugates induced a lower production of anti-native uricase antibodies. The rank order of immunogenicity was native uricase > uricase-poly(N-vinylpyrrolidone) > or = uricase-poly(N-acriloylmorpholine) > uricase-monomethoxypoly(ethylene glycol) 5000 Da > uricase-monomethoxypoly(ethylene glycol) 10000 Da. The four conjugates also induced anti polymer immunoresponse. Anti poly(N-vinylpyrrolidone) and anti poly(N-acriloylmorpholine) antibodies were generated from the first immunization while low levels of anti polymer antibodies were found with both poly(ethylene glycol) conjugates only after the second immunization.     

The effect of homologos amini acid replacement on the conformation of oligopeptides. III. CD studies on co-oligopeptides of methionine and valine or methionine and glycine in organic solution

A conformational analyis of co-ologopeptides containing methionine and valine or methionine and glycine was carried out using circular dichrosim. The oligopeptides containing valine and methionine (dimer to hexamer) are disorder in hexafluoropropane diol·0.5 H₂O and trimethyl phospate but become helical in trifluoroethanol at the heptamer. The CD spectra for hesamers and heptamers containing methionine or methionine and one valine give evidience that one valyl residue can be inserted into these peptides wothout affecting their secondry structure. Co-oligomethionines. The effect of a glycyl residue are generally less ordered than the analogous homo-oligomethionines. The effect of a glycl residue on the structure of the longer oligopeptides depends on its position in the chain. When inserted in the center of a hexamer or heptamer, the single glycyl residue destabilizes the ordered secondry structures in solution. Finally, evidence is presented that the CD patterns observed for various pentamers and hexamers are consistent with some order at these chain lenghts.     

Biochemical problems of regulation by oligopeptides

Some problems are considered which arise in biochemical studies on structure and function of natural oligopeptides consisting of 2–50 amino acid residues. The problems under consideration include the generation of oligopeptides from precursors, chemical structure, the role of functionally important radicals and spatial configuration, and structure-function relationships. Different types of regulation are shown mainly for oligopeptides involved in muscle contraction.Translated from Biokhimiya, Vol. 69, No. 11, 2004, pp. 1565–1573.Original Russian Text Copyright © 2004 by Zamyatnin.     

Hemoglobin as a potential source of natural regulatory oligopeptides

Theoretical structure-function analysis of all possible hemoglobin molecule fragments was performed to determine sites that could be potential sources of regulatory oligopeptides. Known data on bovine hemoglobin primary structure and information of the EROP-Moscow database concerning structure and functions of natural oligopeptides were used along with a computer program complex. A total of 6750 natural non-hemoglobin oligopeptides with hemoglobin fragments of 2–14 amino acid residues were found. Structures of 20 of them were completely identical to hemoglobin fragments. Most of the revealed oligopeptides exhibit properties of neuropeptides, antimicrobial agents, and hormones. A number of them exhibit functions previously not known for hemoglobin fragments. The possibility of natural formation of regulatory oligopeptides from hemoglobin and other food protein molecules, generation of the exogenous oligopeptide pool, their participation in regulation processes as well as accordance of results obtained here with the oligopeptide continuum concepts are discussed.     

Mass spectrometry of acetylated,permethylated and reduced oligopeptides

The amino acid sequence of oligopeptides can be determined from the mass spectra of their volatile derivatives. New peptide derivatives are described which are prepared from permethylated peptides by reduction with borane-tetrahydrofuran. These derivatives have been found to be more volatile than the corresponding permethylated ones. The procedure is illustrated by application of this technique to representative oligopeptides. As little as 10–100 nmol of these peptides have been sequenced using this technique.     

Conformational studies on sequential polypeptides Part VI. Structural investigations on (Pro-Leu-Gly)10, (Pro-Leu-Gly)n and (Leu-Pro-Gly)n.

The conformational properties of (Pro-Leu-Gly)10, (Pro-Leu-Gly)n and (Leu-Pro-Gly)n were investigated both in solution and in solid state. By circular dichroism studies it was possible to demonstrate the formation of an ordered collagen-like structure for (Pro-Leu-Gly)n in hexafluroisopropanol-water mixtures and in ethylene glycol; (Leu-Pro-Gly)n assumes an ordered conformation only in ethylene glycol; (Pro-Leu-Gly)10 is unordered under all the conditions studied. X-ray diffraction patterns indicated that (Pro-Leu-Gly)n and (Leu-Pro-Gly)n assume a triple helical structure in solid state. In addition, the investigation of (Pro-Leu-Gly)n strongly suggests that this type of structure is a single chain triple helix. The X-ray patterns of (Pro-Leu-Gly)10 do not allow us to ascertain a collagen or polyproline II-like structure for this decatripeptide.     

Cationic oligopeptides with the repeating sequence L‐lysyl‐L‐alanyl‐L‐alanine: conformational and thermal stability study using optical spectroscopic methods

The infrared (IR), vibrational circular dichroism (VCD), and electronic circular dichroism (ECD) spectra of short cationic sequential peptides (L ‐Lys‐L ‐Ala‐L ‐Ala)_n (n = 1, 2, and 3) were measured over a range of temperatures (20–90 °C) in aqueous solution at near‐neutral pH values in order to investigate their solution conformations and thermally induced conformational changes. VCD spectra of all three oligopeptides measured in the amide I′ region indicate the presence of extended helical polyproline II (PPII)‐like conformation at room temperature. UV‐ECD spectra confirmed this conclusion. Thus, the oligopeptides adopt a PPII‐like conformation, independent of the length of the peptide chain. However, the optimized dihedral angles ? and ψ are within the range ?82 to ?107° and 143–154°, respectively, and differ from the canonical PPII values. At elevated temperatures, the observed intensity and bandshape variations in the VCD and ECD spectra show that the PPII‐like conformation of the Lys‐Ala‐Ala sequence is still preferred, being in equilibrium with an unordered conformer at near‐neutral pH values within the range of temperatures from 20 to 90 °C. This finding was obtained from analysis of the temperature‐dependent spectra using the singular value decomposition method. The study presents KAA‐containing oligopeptides as conformationally stable models of biologically important cationic peptides and proteins. Copyright © 2009 European Peptide Society and John Wiley & Sons, Ltd.     

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.     

Homochiral oligopeptides via surface recognition and enantiomeric cross impediment in the polymerization of racemic phenylalanine N-carboxyanhydride crystals suspended in water

As part of our program on the search of possible prebiotic routes for the formation of oligopeptides of homochiral sequence (isotactic) from racemic precursors in aqueous environment, we report the polymerization of racemic crystals of phenylalanine N-carboxyanhydrides, enantioselectively tagged with five deuterium atoms, suspended in water containing various amine initiators. Racemic mixtures of isotactic oligopeptides, comprising up to 25 repeat units of the same handedness, as the dominant component for each length, were observed in a MALDI-TOF mass spectrometry analysis. The racemic mixtures of the peptides could be desymmetrized by initiating the polymerization reaction with water-soluble methyl esters of either enantiopure alpha-amino acids or dipeptides. A three-step mechanism is proposed to account for these results: (i) Surface recognition of the chiral initiator by the chiral sites present at specific faces of the crystal; (ii) Oligopeptide elongation at the polymer/crystal interface; and (iii) Self-assembly of the short isotactic peptides into racemic antiparallel beta-sheets as templates followed by cross-enantiomeric impediment in the growth of enantiomeric chains at the peptide beta-sheet/crystal interface.     

Antimicrobial and other oligopeptides of grapes

Structures and functions of about 700 oligopeptides of various plants are presently known. However, only one polypeptide has been isolated from grapes and characterized. At the same time, tens of thousands of uncharacterized amino acid sequences have been revealed in this plant, among which there can also be precursors of oligopeptide regulators. Due to the scientific and practical importance of innate immunity of agricultural plants, we have undertaken structural and functional investigation of these sequences to identify new regulatory oligopeptides including antimicrobial agents. For this purpose, we elaborated a special method of computer analysis enabling comparison of primary structures of putative precursors of grape oligopeptides with amino acid sequences of known oligopeptides of other plants. Structural similarity served as the basis for prediction of potential functional properties. As a result, over 20 new structures of antimicrobial and other grape oligopeptides have been found.     

Biological activities of retro and diastereo analogs of a 13-residue peptide with antimicrobial and hemolytic activities.

The biological activities of synthetic retro and diastereo analogs of PKLLKTFLSKWIG (SPFK), a 13-residue peptide with antimicrobial and hemolytic activities, have been investigated. Retro peptides with C-terminal acid and amide exhibited antibacterial activities comparable with those of SPFK. Their hemolytic activities were, however, only marginally lower. The diastereo analog with C-terminal acid was not antibacterial and was weakly hemolytic. Amidation of this analog could restore antibacterial activity. Both retro analogs were unordered in aqueous medium but had a propensity for a helical structure in trifluoroethanol. However, diastereo analogs were unordered in both aqueous medium and trifluoroethanol. Thus, reversing the sequence in a short amphiphilic peptide may not always result in the selective loss of biological activity such as hemolytic activity. Also, introduction of enantiomeric amino acids in a short peptide to generate a diastereomer may result in loss of structure as well as antimicrobial and hemolytic activities, unless compensated by an increase in positive charges.     

The interaction of phospholipid membranes with poly(ethylene glycol). Vesicle aggregation and lipid exchange

(1) The water soluble polymer, poly(ethylene glycol), causes aggregation of sonicated vesicles of dimyristoylphosphatidylcholine in a manner consistent with a steric exclusion mechanism. (2) Poly(ethylene glycol) promotes the exchange of lipids between multilamellar vesicles of dimyristoylphosphatidylcholine and dipalmitoylphosphatidylcholine when the lipids are in the liquid-crystalline state. (3) ³¹P-NMR studies demonstrate that the bilayer configuration of smectic mesophases of dipalmitoylphosphatidylcholine is substantially maintained in the presence of poly(ethylene glycol).     

Approach to studying natural oligopeptides in the area of structure relationship to function]

Structure and structure-function relations of naturally occurring oligopeptides and peptide receptors are discussed. An approach to inferring function of low-molecular peptides in the direction from their structure is postulated. Diverse biological activities of oligopeptides supposedly arise from a limited number of preferable spatial structures which may exist under physiological conditions. Each particular function of an oligopeptide is connected with a definite spatial structure, belonging to the set of the low-energy conformations. A method is suggested for constructing a synthetic analogue with a predetermined physiologically active conformation, prior to all chemical and biological tests.