Home About us Contact
|
Home About us Contact | ||
|
|
||
Small Peptides (small + peptide)
Selected AbstractsSmall peptides, big world: biotechnological potential in neglected bioactive peptides from arthropod venoms,JOURNAL OF PEPTIDE SCIENCE, Issue 11 2005Adriano M. C. Pimenta Abstract Until recently, a toxinologist's tasks involved the search for highly toxic or lethal toxins in animal venoms that could explain the harmful effects in clinically observed symptoms. Most of these toxins were put on evidence using a function to structure approach, in which a biological phenomena observation usually guided the isolation and characterization of the causative molecule. Paving this way, many toxins were promptly purified because of their readily observed effect. Nevertheless, small molecules with micro-effects that are not easily visualized can be relatively neglected or poorly studied. This situation has changed now with the advent of the sensitivity, resolution and accuracy of techniques such as mass spectrometry and proteomic approaches used in toxinology. Taking advantage of these methodologies, small peptides with ,newly exploited' biological activities such as vasoactive, hormone-like, antimicrobial and others have been recently given much more attention, enlarging the known repertoire of bioactive molecules found in animal venoms. This article aims to review current knowledge on small biologically active peptides (<3 kDa) found in arthropod venoms and discuss their potentialities as new drug candidates or therapeutic lead compounds. Copyright © 2005 European Peptide Society and John Wiley & Sons, Ltd. [source] Adventures in multivalency, the Harry S. Fischer memorial lecture CMR 2005; Evian, FranceCONTRAST MEDIA & MOLECULAR IMAGING, Issue 1 2006Michael F. Tweedle Abstract This review discusses multivalency in the context of drug discovery, specifically the discovery of new diagnostic imaging and related agents. The aim is to draw attention to the powerful role that multivalency plays throughout research involving molecular biology, in general, and much of biochemically targeted contrast agent research, in particular. Two examples from the author's laboratory are described. We created small (,5,kDa) peptide ,dimers' composed of two different, chemically linked peptides. The monomer peptides both bound to the same target protein with Kd,,,100,s,nM, while the heterodimers had sub-nM Kd values. Biological activity was evident in the heterodimers where none or very little existed in homodimers, monomers or monomer mixtures. Two different tyrosine kinases (KDR and C-Met) and four peptide families produced consistent results: multivalent heterodimers were uniquely different. The second example begins with making two micron ultrasound bubbles coated with the peptide, TKPPR (a Tuftsin antagonist) as a negative control for bubbles targeted with angiogenesis target-binding peptides. Unexpected binding of a ,negative' control, (TKPPR)-targeted bubble to endothelial cells expressing angiogenesis targets, led to the surprising result that TKPPR, only when multimerized, binds avidly, specifically and actively to neuropilin-1, a VEGF co-receptor. VEGF is the primary stimulator of angiogenesis. Tuftsin is a small peptide (TKPR) derived from IgG that binds to macrophages during inflammation, and has been studied for over 30 years. The receptor has never been cloned. The results led to new conclusions about Tuftsin, neuropilin-1 and the purpose, up to now unknown, of exon 8 in VEGF. Multivalency can be used rationally to solve practical problems in drug discovery. When targeting larger structures, multivalency is frequently unavoidable, and can lead to unpredictable and useful biochemical information, as well as to new drug candidates. Copyright © 2006 John Wiley & Sons, Ltd. [source] Hepcidin , central regulator of iron metabolismEUROPEAN JOURNAL OF HAEMATOLOGY, Issue 1 2007Valeriu Atanasiu Abstract The knowledge about mammalian iron metabolism has advanced dramatically over the past decades. Studies of genetics, biochemistry and molecular biology allowed us the identification and characterization of many of the molecules involved in regulation of iron homeostasis. Important progresses were made after the discovery in 2000 of a small peptide , hepcidin , that has been proved to play a central role in orchestration on iron metabolism also providing a link between iron metabolism and inflammation and innate immunity. Hepcidin directly interacts with ferroportin (FPN), the only known mammalian iron exporter, which is expressed by enterocytes, macrophages and hepatocytes. The direct hepcidin,FPN interaction allows an adaptative response from the body in situations that alter normal iron homeostasis (hypoxia, anemia, iron deficiency, iron overload, and inflammation). [source] Synthesis, Solution Structure and Biological Activity of Val-Val-Pro-Gln,a Bioactive Elastin PeptideEUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 8 2005Caterina Spezzacatena Abstract Val-Val-Pro-Gln (valyl-valyl-prolyl-glutamine) is a small but highly conserved sequence present in all elastins. We describe its synthesis by mixed anhydride solution chemistry as an alternative to solid-phase peptide synthesis (SPPS). The molecular structure of the tetrapeptide in solution was investigated by classical spectroscopy, such as circular dichroism (CD), nuclear magnetic resonance (NMR) and Fourier Transform Infrared Spectroscopy (FTIR). The biological activity of Val-Val-Pro-Gln was evaluated by a bromodeoxyuridine (BrdU) incorporation assay with normal human dermal fibroblasts. This small peptide may play a critical role in control of matrix metabolism through its release from the elastin polypeptide chain during periods of tissue breakdown and remodelling. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005) [source] C-terminal 37 residues of LRP promote the amyloidogenic processing of APP independent of FE65JOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 6b 2008Madepalli K. Lakshmana Abstract The major defining pathological hallmark of Alzheimer's disease (AD) is the accumulation of amyloid , protein (A,), a small peptide derived from ,- and ,-secretase cleavages of the amyloid precursor protein (APP). Recent studies have shown that the Low-density lipoprotein receptor-related protein (LRP) plays a pivotal role in the trafficking of APP and generation of A,. In particular, we recently showed that the soluble cytoplasmic tail of LRP (LRP-ST) without a membrane tether was sufficient to promote A, generation. In this study, we demonstrate that the last 37 residues of LRP cytoplasmic tail (LRP-C37) lacking the NPxY motifs and FE65 binding mediate the core pro-amyloidogenic activity of LRP-ST. Moreover, we show that the conserved dileucine motif within the LRP-C37 region is a key determinant of its A, promoting activity. Finally, results from a yeast two-hybrid screen using LRP-C37 region as bait reveal four new LRP-binding proteins implicated in intracellular signalling and membrane protein trafficking. Our findings indicate that the LRP-C37 sequence represents a new protein-binding domain that may be useful as a therapeutic target and tool to lower A, generation in AD. [source] Recombinant vascular basement membrane derived multifunctional peptide blocks endothelial cell angiogenesis and neovascularization,JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2010Chengkun Wang Abstract Angiogenesis is an innovative target in the therapy of cancer and other diseases, but the effects of anti-angiogenic drugs have been rather modest in clinical trials. We have developed a small peptide, recombinant vascular basement membrane derived multifunctional peptide (rVBMDMP), which significantly inhibits endothelial cells in vitro. Here we test the mechanisms of rVBMDMP in angiogenesis balance in assays of tubule formation, colony formation, and apoptosis in HUVE-12 endothelial cells. We also analyzed the differential expression of phosphorylation proteins and related genes in a protein phosphorylation chip and extracellular matrix adhesion molecule cDNA microarray, and validated changes with Western blot or real-time quantitative PCR, respectively. rVBMDMP dose-dependently inhibited colony formation, induced apoptosis, and inhibited in vitro tubule formation. rVBMDMP increased the phosphorylation of 88 signal proteins, including caspase-3, death receptor 3, 4, and 5, and integrin ,V, ,1, and ,3, and down-regulated 41 signal proteins, including EGFR, pEGFR, VEGFR-1, and survivin versus control. rVBMDMP upregulated 14 genes, including collagen 4, 7, and 27, and down-regulated 21 genes, including integrin ,V,3, MMP10, and MMP12. Our study suggests that rVBMDMP inhibits angiogenesis and may be a viable drug candidate in anti-angiogenesis and anticancer therapies. J. Cell. Biochem. 111: 453,460, 2010. © 2010 Wiley-Liss, Inc. [source] Improving the affinity of antigens for mutated antibodies by use of statistical molecular designJOURNAL OF PEPTIDE SCIENCE, Issue 7 2008Ilona Mandrika Abstract We demonstrate the use of statistical molecular design (SMD) in the selection of peptide libraries aimed to systematically investigate antigen-antibody binding spaces. Earlier, we derived two novel antibodies by mutating the complementarity-determining region of the anti-p24 (HIV-1) single chain Fv antibody, CB4-1 that had lost their affinity for a p24 epitope-homologous peptide by 8- and 60-fold. The present study was devoted to explore how peptide libraries can be designed under experimental design criteria for effective screening of peptide antigens. Several small peptide,antigen libraries were selected using SMD principles and their activities were evaluated by their binding to SPOT-synthesized peptide membranes and by fluorescence polarization (FP). The approach was able to reveal the most critical residues required for antigen binding, and finally to increase the binding activity by proper modifications of amino acids in the peptide antigen. A model of the active peptide binding pocket formed by the mutated scFv and the antigen was compatible with the information gained from the experimental data. Our results suggest that SMD approaches can be used to explore peptide antigen features essential for their interactions with antibodies. Copyright © 2008 European Peptide Society and John Wiley & Sons, Ltd. [source] The cia operon of Streptococcus mutans encodes a unique component required for calcium-mediated autoregulationMOLECULAR MICROBIOLOGY, Issue 1 2008Xuesong He Summary Streptococcus mutans is a primary pathogen for dental caries in humans. CiaR and CiaH of S. mutans comprise a two-component signal transduction system (TCS) involved in regulating various virulent factors. However, the signal that triggers the CiaRH response remains unknown. In this study, we show that calcium is a signal for regulation of the ciaRH operon, and that a double-glycine-containing small peptide encoded within the ciaRH operon (renamed ciaX) mediates this regulation. CiaX contains a serine + aspartate (SD) domain that is shared by calcium-binding proteins. A markerless in-frame deletion of ciaX reduced ciaRH operon expression and diminished the calcium repression of operon transcription. Point mutations of the SD domain resulted in the same phenotype as the in-frame deletion, indicating that the SD domain is required for CiaX function. Further characterization of ciaX demonstrated that it is involved in calcium-mediated biofilm formation. Furthermore, inactivation of ciaR or ciaH led to the same phenotype as the in-frame deletion of ciaX, suggesting that all three genes are involved in the same regulatory pathway. Sequence analysis and real-time RT-PCR identified a putative CiaR binding site upstream of ciaX. We conclude that the ciaXRH operon is a three-component, self-regulatory system modulating cellular functions in response to calcium. [source] How do helix,helix interactions help determine the folds of membrane proteins?PROTEIN SCIENCE, Issue 4 2003Perspectives from the study of homo-oligomeric helical bundles FRET, fluorescence resonance energy transfer; NBD, 7-nitrobenz-2-oxa-1,3-diazole; C-14 betaine, N -tetradecyl- N,N -dimethyl-3-ammonio-1-propanesulfonate; MF, mole fraction Abstract The final, structure-determining step in the folding of membrane proteins involves the coalescence of preformed transmembrane helices to form the native tertiary structure. Here, we review recent studies on small peptide and protein systems that are providing quantitative data on the interactions that drive this process. Gel electrophoresis, analytical ultracentrifugation, and fluorescence resonance energy transfer (FRET) are useful methods for examining the assembly of homo-oligomeric transmembrane helical proteins. These methods have been used to study the assembly of the M2 proton channel from influenza A virus, glycophorin, phospholamban, and several designed membrane proteins,all of which have a single transmembrane helix that is sufficient for association into a transmembrane helical bundle. These systems are being studied to determine the relative thermodynamic contributions of van der Waals interactions, conformational entropy, and polar interactions in the stabilization of membrane proteins. Although the database of thermodynamic information is not yet large, a few generalities are beginning to emerge concerning the energetic differences between membrane and water-soluble proteins: the packing of apolar side chains in the interior of helical membrane proteins plays a smaller, but nevertheless significant, role in stabilizing their structure. Polar, hydrogen-bonded interactions occur less frequently, but, nevertheless, they often provide a strong driving force for folding helix,helix pairs in membrane proteins. These studies are laying the groundwork for the design of sequence motifs that dictate the association of membrane helices. [source] On the application of an experimental multipolar pseudo-atom library for accurate refinement of small-molecule and protein crystal structuresACTA CRYSTALLOGRAPHICA SECTION A, Issue 2 2007Bartosz Zarychta With an increasing number of biomacromolecular crystal structures being measured to ultra-high resolution, it has become possible to extend to large systems experimental charge-density methods that are usually applied to small molecules. A library has been built of average multipole populations describing the electron density of chemical groups in all 20 amino acids found in proteins. The library uses the Hansen & Coppens multipolar pseudo-atom model to derive molecular electron density and electrostatic potential distributions. The library values are obtained from several small peptide or amino acid crystal structures refined against ultra-high-resolution X-ray diffraction data. The library transfer is applied automatically in the MoPro software suite to peptide and protein structures measured at atomic resolution. The transferred multipolar parameters are kept fixed while the positional and thermal parameters are refined. This enables a proper deconvolution of thermal motion and valence-electron-density redistributions, even when the diffraction data do not extend to subatomic resolution. The use of the experimental library multipolar atom model (ELMAM) also has a major impact on crystallographic structure modelling in the case of small-molecule crystals at atomic resolution. Compared to a spherical-atom model, the library transfer results in a more accurate crystal structure, notably in terms of thermal displacement parameters and bond distances involving H atoms. Upon transfer, crystallographic statistics of fit are improved, particularly free R factors, and residual electron-density maps are cleaner. [source] A comparison between experimental and theoretical aspherical-atom scattering factors for charge-density refinement of large moleculesACTA CRYSTALLOGRAPHICA SECTION A, Issue 3 2004Virginie Pichon-Pesme The differences between two databases describing the polypeptide main chain in terms of charge-density parameters, directly usable in protein structure refinements, are discussed. These databases contain averaged multipole populations of peptide pseudo-atoms obtained from refinement against theoretical simulated data and against high-resolution experimental data on small peptide or amino acid molecules. The main discrepancy becomes apparent when electrostatic properties are calculated. [source] Structure of the twin-arginine signal-binding protein DmsD from Escherichia coliACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2009Suresh Kumar Ramasamy The translocation of folded proteins via the twin-arginine translocation (Tat) pathway is regulated to prevent the futile export of inactive substrate. DmsD is part of a class of cytoplasmic chaperones that play a role in preventing certain redox proteins from premature transport. DmsD from Escherichia coli has been crystallized in space group P41212, with unit-cell parameters a = b = 97.45, c = 210.04,Å, in the presence of a small peptide. The structure has been solved by molecular replacement to a resolution of 2.4,Å and refined to an R factor of 19.4%. There are four molecules in the asymmetric unit that may mimic a higher order structure in vivo. There appears to be density for the peptide in a predicted binding pocket, which lends support to its role as the signal-recognition surface for this class of proteins. [source] Identification and Repair of Positive Binding Antibodies Containing Randomly Generated Amber Codons from Synthetic Phage Display LibrariesBIOTECHNOLOGY PROGRESS, Issue 3 2006Warren D. Marcus Phage display technology allows for the rapid isolation and characterization of monoclonal antibodies that have vast potential for therapeutic and diagnostic applications. However, the panning process, which utilizes a host strain that suppresses termination by the amber codon, has an inherent bias toward clones containing randomly generated amber stop codons, complicating identification of positive binding antibodies when the antibody genes are finally expressed in a nonsupressor host. Here, we perform biopanning against a Histone 2A peptide using streptavidin- or anti-biotin-coated beads. After four rounds, a dominant clone is characterized but contains a spurious amber stop codon. A protocol is given that readily corrects the amber codon, allowing for soluble antibody production once the phagemid is transformed into a nonsuppressor bacterial strain. This work also highlights the ability to isolate antibodies against a protein antigen by using only a small peptide (15 amino acids) representing a portion of the antigen. [source] Cell Cycle Progression in Serum-Free Cultures of Sf9 Insect Cells: Modulation by Conditioned Medium Factors and Implications for Proliferation and Productivity,BIOTECHNOLOGY PROGRESS, Issue 5 2000Magnus Doverskog Cell cycle progression was studied in serum-free batch cultures of Spodoptera frugiperda (Sf9) insect cells, and the implications for proliferation and productivity were investigated. Cell cycle dynamics in KBM10 serum-free medium was characterized by an accumulation of 50,70% of the cells in the G2/M phase of the cell cycle during the first 24 h after inoculation. Following the cell cycle arrest, the cell population was redistributed into G1 and in particular into the S phase. Maximum rate of proliferation (,N,max) was reached 24,48 h after the release from cell cycle arrest, coinciding with a minimum distribution of cells in the G2/M phase. The following declining ,N could be explained by a slow increase in the G2/M cell population. However, at approximately 100 h, an abrupt increase in the amount of G2/M cells occurred. This switch occurred at about the same time point and cell density, irrespective of medium composition and maximum cell density. An octaploid population evolved from G2/M arrested cells, showing the occurrence of endoreplication in this cell line. In addition, conditioned medium factor(s) were found to increase ,N,max, decrease the time to reach ,N,max, and decrease the synchronization of cells in G2/M during the lag and growth phase. A conditioned medium factor appears to be a small peptide. On basis of these results we suggest that the observed cell cycle dynamics is the result of autoregulatory events occurring at key points during the course of a culture, and that entry into mitosis is the target for regulation. Infecting the Sf9 cells with recombinant baculovirus resulted in a linear increase in volumetric productivity of ,-galactosidase up to 68,75 h of culture. Beyond this point almost no product was formed. Medium renewal at the time of infection could only partly restore the lost hypertrophy and product yield of cultures infected after the transition point. The critical time of infection correlated to the time when the mean population cell volume had attained a minimum, and this occurred 24 h before the switch into the G2/M phase. We suggest that the cell density dependent decrease in productivity ultimately depends on the autoregulatory events leading to G2/M cell cycle arrest. [source] Pseudo 3D Single-Walled Carbon Nanotube Film for BSA-Free Protein ChipsCHEMBIOCHEM, Issue 8 2005Hye Ryung Byon Sweet home for proteins. Single-walled carbon nanotube (SWNT) film treated with CDI,Tween20 (CT) has been used as a substrate for a BSA-free protein chip. In the absence of BSA, which is conventionally applied as a nonspecific binding inhibitor, protein A, biotinylated BSA, and even small peptide such as 3×FLAG immobilized on the CT-functionalized SWNT film substrates recognize their respective counterparts, streptavidin, immunoglobulin G, and antiFLAG. with high specificity. [source] Site-directed PEGylations of Thymosin , 1 Analogs and Evaluation of Their ImmunoactivityCHINESE JOURNAL OF CHEMISTRY, Issue 4 2009Jiankun QIE Abstract PEGylation is an effective way to improve the pharmacokinetic profiles of pharmaceutical proteins or peptides. But the relatively large and long PEG chains would be likely to shelter the active site of a small peptide because of its small size, compared with a protein. Therefore, the positions and numbers of PEGylation are crucial for the bioactivity of a PEGylated peptide. To elucidate the relationship between the PEGylated positions and bioactivity of a peptide drug, site-specific PEGylations were performed on Zadaxin (Thymosin , 1, T,1), which is a pharmaceutical peptide with an , -helix region, a , -turn region, and random coils. Site-specific mono-PEGylations of T,1 in different conformational regions were realized through introducing one cysteine residue into the desired positions of the peptide, followed by a coupling reaction with a thiol-attached maleimide-PEG reagent. Primary data from IFN- , production of splenocytes induced by Con A showed that the influence of PEGylation on Zadaxin was position-dependent, and mostly, positive effects were observed after PEGylation, which indicated that the position of PEGylation is important for maintaining the bioactivity of a peptide. [source] Voltage-gated sodium channel isoform-specific effects of pompilidotoxinsFEBS JOURNAL, Issue 4 2010Emanuele Schiavon Pompilidotoxins (PMTXs, , and ,) are small peptides consisting of 13 amino acids purified from the venom of the solitary wasps Anoplius samariensis (,-PMTX) and Batozonellus maculifrons (,-PMTX). They are known to facilitate synaptic transmission in the lobster neuromuscular junction, and to slow sodium channel inactivation. By using ,-PMTX, ,-PMTX and four synthetic analogs with amino acid changes, we conducted a thorough study of the effects of PMTXs on sodium current inactivation in seven mammalian voltage-gated sodium channel (VGSC) isoforms and one insect VGSC (DmNav1). By evaluating three components of which the inactivating current is composed (fast, slow and steady-state components), we could distinguish three distinct groups of PMTX effects. The first group concerned the insect and Nav1.6 channels, which showed a large increase in the steady-state current component without any increase in the slow component. Moreover, the dose-dependent increase in this steady-state component was correlated with the dose-dependent decrease in the fast component. A second group of effects concerned the Nav1.1, Nav1.2, Nav1.3 and Nav1.7 isoforms, which responded with a large increase in the slow component, and showed only a small steady-state component. As with the first group of effects, the slow component was dose-dependent and correlated with the decrease in the fast component. Finally, a third group of effects concerned Nav1.4 and Nav1.5, which did not show any change in the slow or steady-state component. These data shed light on the complex and intriguing behavior of VGSCs in response to PMTXs, helping us to better understand the molecular determinants explaining isoform-specific effects. [source] Characterization of membrane-bound prolyl endopeptidase from brainFEBS JOURNAL, Issue 17 2008Jofre Tenorio-Laranga Prolyl oligopeptidase (POP) is a serine protease that cleaves small peptides at the carboxyl side of an internal proline residue. Substance P, arginine,vasopressin, thyroliberin and gonadoliberin are proposed physiological substrates of this protease. POP has been implicated in a variety of brain processes, including learning, memory, and mood regulation, as well as in pathologies such as neurodegeneration, hypertension, and psychiatric disorders. Although POP has been considered to be a soluble cytoplasmic peptidase, significant levels of activity have been detected in membranes and in extracellular fluids such as serum, cerebrospinal fluid, seminal fluid, and urine, suggesting the existence of noncytoplasmic forms. Furthermore, a closely associated membrane prolyl endopeptidase (PE) activity has been previously detected in synaptosomes and shown to be different from the cytoplasmic POP activity. Here we isolated, purified and characterized this membrane-bound PE, herein referred to as mPOP. Although, when attached to membranes, mPOP presents certain features that distinguish it from the classical POP, our results indicate that this protein has the same amino acid sequence as POP except for the possible addition of a hydrophobic membrane anchor. The kinetic properties of detergent-soluble mPOP are fully comparable to those of POP; however, when attached to the membranes in its natural conformation, mPOP is significantly less active and, moreover, it migrates anomalously in SDS/PAGE. Our results are the first to show that membrane-bound and cytoplasmic POP are encoded by variants of the same gene. [source] Insect silk contains both a Kunitz-type and a unique Kazal-type proteinase inhibitorFEBS JOURNAL, Issue 7 2001Xavier Nirmala Insect silk is made up of structural fibrous (fibroins) and sticky (sericins) proteins, and contains a few small peptides of hitherto unknown functions. We demonstrate that two of these peptides inhibit bacterial and fungal proteinases (subtilisin, proteinase K and pronase). These ,silk proteinase inhibitors' 1 and 2 (SPI 1 and 2) are produced in the middle section of the silk-secreting glands prior to cocoon spinning and their production is controlled at transcription level. The full length cDNA of pre-SPI 1 contains 443 nucleotides and encodes a peptide of 76 amino-acid residues, of which 20 make up a signal sequence. The mature SPI 1 (6056.7 Da, 56 residues) is a typical thermostable Kunitz-type proteinase inhibitor with Arg in P1 position. The cDNA of pre-SPI 2 consists of 260 nucleotides and yields a putative secretory peptide of 58 amino-acid residues. The functional SPI 2 (3993 Da, 36 residues) is a single-domain Kazal-type proteinase inhibitor with unique structural features: free segment of the N-terminus is reduced to a single amino-acid residue, lack of CysI and CysV precludes formation of the A-ring and provides increased flexibility to the C-ring, and absence of several residues around the normal position of CysV shortens and changes the , helix segment of the protein. The structure reveals that the length and arrangement of the B-ring, including exposure of the P1 residue, and the position of the C-terminus relative to the B-loop, are essential for the activity of the Kazal-type inhibitors. [source] Whey-derived free fatty acids suppress the germination of Candida albicans in vitroFEMS YEAST RESEARCH, Issue 2 2007Martin Clément Abstract Bovine whey from the cheese-making industry contains several bioactive factors that promote health and prevent disease. Although many efforts have been made over the years to show that immunoglobulins, lactoperoxidase, lactoferrin, lysosyme and small peptides present in whey have antimicrobial activities against several pathogenic microorganisms, such activities have not been investigated so far for the lipid fraction of whey. Here, we have used an in vitro assay-based fractionation procedure to show that free fatty acids derived from whey cream specifically inhibit the germination of Candida albicans, a morphologic change associated with pathogenicity. Further fractionation by HPLC demonstrated that this activity can be mainly attributed to lauric acid, myristoleic acid, linoleic acid and arachidonic acid. [source] In vitro haem solubility of red cell fraction of porcine blood under various treatmentsINTERNATIONAL JOURNAL OF FOOD SCIENCE & TECHNOLOGY, Issue 4 2010Tong-Xun Liu Summary An in vitro peptic digestion at gastric pH 2.0 was used to assess the haem solubility of red cell fraction (RCF) of porcine blood derived samples. The in vitro haem solubility of the RCF decreased greatly in the denaturated form of haemoglobin. However, the native haemoglobin was susceptible to be hydrolysed by enzyme mixture of Flavourzyme and Alcalase. The in vitro digestion of the hydrolysates showed that the highest haem solubility was reached at degree of hydrolysis between 8.75% and 12.33%. The in vitro haem solubility was positively correlated with content of the highly soluble peptides with molecular weights ranged from 7.5 kDa to 1 kDa, but negatively with peptides fractions >7.5 kDa and <1 kDa, mostly due to the precipitation of the highly molecular weight fraction (>7.5 kDa) and part of small peptides (<1 kDa) with higher haem/peptide ratio, which was confirmed by gel filtration chromatograms and by the analysis of the precipitate at pH 2.0. [source] The search for low energy conformational families of small peptides: Searching for active conformations of small peptides in the absence of a known receptor,INTERNATIONAL JOURNAL OF QUANTUM CHEMISTRY, Issue 15 2007Katrina W. Lexa Abstract Breast cancer is the most common cancer among women. Tamoxifen is the preferred drug for estrogen receptor-positive breast cancer treatment, yet many of these cancers are intrinsically resistant to tamoxifen or acquire resistance during treatment. Therefore, scientists are searching for breast cancer drugs that have different molecular targets. Previous work revealed that 8-mer and cyclic 9-mer peptides inhibit breast cancer in mouse and rat model systems, interacting with an unknown receptor, while peptides smaller than eight amino acids did not inhibit breast cancer. We have shown that the use of replica exchange molecular dynamics predicts structure and dynamics of active peptides, leading to the discovery of smaller peptides with full biological activity. These simulations identified smaller peptide analogs with a conserved turn, a ,-turn formed in the larger peptides. These analogs inhibit estrogen-dependent cell growth in a mouse uterine growth assay, a test showing reliable correlation with human breast cancer inhibition. We outline the computational methods that were tried and used with the experimental information that led to the successful completion of this research. © 2007 Wiley Periodicals, Inc. Int J Quantum Chem, 2007 [source] Time-averaged predictions of folded and misfolded peptides using a reduced physicochemical modelJOURNAL OF COMPUTATIONAL CHEMISTRY, Issue 7 2008Oliver J. Clarke Abstract Energy-based methods for calculating time-averaged peptide structures are important for rational peptide design, for defining local structure propensities in large protein chains, and for exploring the sequence determinants of amyloid formation. High-end methods are currently too slow to be practicable, and will remain so for the foreseeable future. The challenge is to create a method that runs quickly on limited computer resources and emulates reality sufficiently well. We have developed a simplified off-lattice protein model, incorporating semi-empirical physicochemical potentials, and combined it with an efficient Monte Carlo method for calculating time-averaged peptide structures. Reasonably accurate predictions are found for a set of small ,-helical and ,-hairpin peptides, and we demonstrate a potential application in measuring local structure propensities in protein chains. Time-averaged structures have also been calculated for a set of small peptides known to form ,-amyloid fibrils. The simulations were of three interacting peptides, and in each case the time-averaged structure describes a three-stranded ,-sheet. The performance of our method in measuring the propensities of small peptides to self-associate into possible prefibrillar species compares favorably with more detailed and CPU-intensive approaches. © 2007 Wiley Periodicals, Inc. J Comput Chem, 2008 [source] Application of torsion angle molecular dynamics for efficient sampling of protein conformationsJOURNAL OF COMPUTATIONAL CHEMISTRY, Issue 15 2005Jianhan Chen Abstract We investigate the application of torsion angle molecular dynamics (TAMD) to augment conformational sampling of peptides and proteins. Interesting conformational changes in proteins mainly involve torsional degrees of freedom. Carrying out molecular dynamics in torsion space does not only explicitly sample the most relevant degrees of freedom, but also allows larger integration time steps with elimination of the bond and angle degrees of freedom. However, the covalent geometry needs to be fixed during internal coordinate dynamics, which can introduce severe distortions to the underlying potential surface in the extensively parameterized modern Cartesian-based protein force fields. A "projection" approach (Katritch et al. J Comput Chem 2003, 24, 254,265) is extended to construct an accurate internal coordinate force field (ICFF) from a source Cartesian force field. Torsion crossterm corrections constructed from local molecular fragments, together with softened van der Waals and electrostatic interactions, are used to recover the potential surface and incorporate implicit bond and angle flexibility. MD simulations of dipeptide models demonstrate that full flexibility in both the backbone ,/, and side chain ,1 angles are virtually restored. The efficacy of TAMD in enhancing conformational sampling is then further examined by folding simulations of small peptides and refinement experiments of protein NMR structures. The results show that an increase of several fold in conformational sampling efficiency can be reliably achieved. The current study also reveals some complicated intrinsic properties of internal coordinate dynamics, beyond energy conservation, that can limit the maximum size of the integration time step and thus the achievable gain in sampling efficiency. © 2005 Wiley Periodicals, Inc. J Comput Chem 26: 1565,1578, 2005 [source] PROTEOLYSIS IN SALMON (SALMO SALAR) DURING COLD STORAGE; EFFECTS OF STORAGE TIME AND SMOKING PROCESSJOURNAL OF FOOD BIOCHEMISTRY, Issue 5 2001KAREN ENGVANG LUND ABSTRACT Changes in free amino acids (FAAs), small peptides and myofibrillar proteins were investigated in salmon (Salmo salar) muscle stored at OC for up to 23 days and after the stored salmon was smoked. Storage time and smoking process did not increase the formation of FAAs and small peptides indicating low exopeptidase activity. During storage, SDS PAGE analysis of myofibrils showed an increase in density of bands at 16, 37, 60, 64, 67, 76 and 130 kDa, a decrease of a 32 kDa band and the appearance of four new bands of 30, 90, 95 and 113 kDa. These results indicate proteolytic degradation. A little change of myosin and no change of ,-actinin and actin were observed. The smoking process itself enhances the intensity of bands, but does not change the pattern markedly except for the appearance of a 25 and 70 kDa band. Degradation pattern after smoking was not affected by storage time. [source] Preparation and Characterization of Hydrolyzed Proteins from Defibrinated Bovine PlasmaJOURNAL OF FOOD SCIENCE, Issue 2 2002P.K.J.P.D. Wanasundara ABSTRACT: Proteins from defibrinated bovine blood plasma were enzymatically hydrolyzed with food-grade microbial proteases Alcalase 2.4 L® and Flavourzyme L&TM;, and a substrate consisting of small peptides and free amino acids was obtained. Hydrolysis of the plasma proteins with Flavourzyme resulted in a maximum degree of hydrolysis of 43% at an enzyme concentration of 110 LAPU/g protein after 15.5 h of hydrolysis. Among the free amino acids in the hydrolysate, hydrophobic amino acids were predominant. The major plasma proteins were degraded due to hydrolysis; peptides of less than 1.04 kDa were dominant in the product when a high degree of hydrolysis was employed. [source] A fast, reproducible and low-cost method for sequence deconvolution of ,on-bead' peptides via ,on-target' maldi-TOF/TOF mass spectrometryJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 3 2010Giulio A. Amadei Abstract A novel approach to high-throughput sequence deconvolution of on-bead small peptides (MW < 2000 Da) using on-target MALDI-TOF/TOF instrumentation is presented. Short peptides of pentamer and octamer length, covalently attached to TentaGel polystyrene beads through a photolabile linker, were placed onto the MALDI target, apportioned with suitable matrix (2,5-dihydroxybenzoic acid) and then hit with the instrument laser (Nd : YAG, 355 nm). This induced easy and highly reproducible photochemical cleavage, desorption (MS mode) and fragmentation (MS/MS mode). Peptide fragments were identified with a mass accuracy of 0.1 Da of the expected values. This technique significantly accelerates the sequence determination of positive peptide hits obtained from random combinatorial libraries when screening against biological targets, paving the way for a rapid and efficient method to identify molecular imaging ligands specific to pathological targets in cancer and other diseases. Copyright © 2009 John Wiley & Sons, Ltd. [source] Peptides derived from the solvent-exposed loops 3 and 4 of BDNF bind TrkB and p75NTR receptors and stimulate neurite outgrowth and survivalJOURNAL OF NEUROSCIENCE RESEARCH, Issue 6 2010Kristina Fobian Abstract Brain-derived neurotrophic factor (BDNF) is critically involved in modeling the developing nervous system and is an important regulator of a variety of crucial functions in the mature CNS. BDNF exerts its action through interactions with two transmembrane receptors, either separately or in concert. BDNF has been implicated in several neurological disorders, and irregularities in BDNF function may have severe consequences. Administration of BDNF as a drug has thus far yielded few practicable results, and the potential side effects when using a multifunctional protein are substantial. In an effort to produce more specific compounds without side effects, small peptides mimicking protein function have been developed. The present study characterized two mimetic peptides, Betrofin 3 and Betrofin 4, derived from the BDNF sequence. Both Betrofins bound the cognate BDNF receptors, TrkB and p75NTR, and induced neurite outgrowth and enhanced neuronal survival, probably by inducing signaling through tha Akt and MAPK pathways. Distinct, charged residues within the Betrofin sequences were identified as important for generating the neuritogenic response, which was also inhibited when BDNF was added together with either Betrofin, indicating partial agonistic effects of the peptides. Thus, two peptides derived from BDNF induced neurite outgrowth and enhanced neuronal survival, probably through binding to BDNF receptors. © 2009 Wiley-Liss, Inc. [source] Identification of small peptides mimicking the R2 C -terminus of Mycobacterium tuberculosis ribonucleotide reductaseJOURNAL OF PEPTIDE SCIENCE, Issue 3 2010Daniel J. Ericsson Abstract Ribonucleotide reductase (RNR) is a viable target for new drugs against the causative agent of tuberculosis, Mycobacterium tuberculosis. Previous work has shown that an N -acetylated heptapeptide based on the C -terminal sequence of the smaller RNR subunit can disrupt the formation of the holoenzyme sufficiently to inhibit its function. Here the synthesis and binding affinity, evaluated by competitive fluorescence polarization, of several truncated and N -protected peptides are described. The protected single-amino acid Fmoc-Trp shows binding affinity comparable to the N -acetylated heptapeptide, making it an attractive candidate for further development of non-peptidic RNR inhibitors. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd. [source] ,Sulfo-click' for ligation as well as for site-specific conjugation with peptides, fluorophores, and metal chelatorsJOURNAL OF PEPTIDE SCIENCE, Issue 1 2010Dirk T. S. Rijkers Abstract The ,sulfo-click' reaction, which is a chemoselective amidation reaction involving the reaction of an aminoethane sulfonyl azide with a thio acid, encompasses a new approach for ligation and conjugation. Detailed protocols are provided for decorating biologically active peptides or dendrimers with biophysical tags, fluorescent probes, metal chelators, and small peptides by using this reaction as a novel, metal-free ,sulfo-click' approach. Copyright © 2009 European Peptide Society and John Wiley & Sons, Ltd. [source] | ||||||||||||||||||||||||||||||||||