Peptide H3 Research Articles

Hempseed (Cannabis sativa) Peptide H3 (IGFLIIWV) Exerts Cholesterol-Lowering Effects in Human Hepatic Cell Line.

Hempseed (Cannabis sativa) protein is an important source of bioactive peptides. H3 (IGFLIIWV), a transepithelial transported intestinal peptide obtained from the hydrolysis of hempseed protein with pepsin, carries out antioxidant and anti-inflammatory activities in HepG2 cells. In this study, the main aim was to assess its hypocholesterolemic effects at a cellular level and the mechanisms behind this health-promoting activity. The results showed that peptide H3 inhibited the 3-hydroxy-3-methylglutaryl co-enzyme A reductase (HMGCoAR) activity in vitro in a dose-dependent manner with an IC50 value of 59 μM. Furthermore, the activation of the sterol regulatory element binding proteins (SREBP)-2 transcription factor, followed by the increase of low-density lipoprotein (LDL) receptor (LDLR) protein levels, was observed in human hepatic HepG2 cells treated with peptide H3 at 25 µM. Meanwhile, peptide H3 regulated the intracellular HMGCoAR activity through the increase of its phosphorylation by the activation of AMP-activated protein kinase (AMPK)-pathways. Consequently, the augmentation of the LDLR localized on the cellular membranes led to the improved ability of HepG2 cells to uptake extracellular LDL with a positive effect on cholesterol levels. Unlike the complete hempseed hydrolysate (HP), peptide H3 can reduce the proprotein convertase subtilisin/kexin 9 (PCSK9) protein levels and its secretion in the extracellular environment via the decrease of hepatic nuclear factor 1-α (HNF1-α). Considering all these evidences, H3 may represent a new bioactive peptide to be used for the development of dietary supplements and/or peptidomimetics for cardiovascular disease (CVD) prevention.

Design and biological testing of peptidic dimerization inhibitors of human Hsp90 that target the C-terminal domain

BackgroundSmall molecules targeting the dimerization interface of the C-terminal domain of Hsp90, a validated target for cancer treatment, have yet to be identified. MethodsThree peptides were designed with the aim to inhibit the dimerization of Hsp90. Computational and biophysical methods examined the α-helical structure for the three peptides. Based on the Autodisplay technology, a novel flow cytometer dimerization assay was developed to test inhibition of Hsp90 dimerization. Microscale thermophoresis was used to determine the KD of the peptides towards the C-terminal domain of Hsp90. ResultsMD simulations and CD spectroscopy indicated an α-helical structure for two of the three peptides. By flow cytometer analysis, IC50 values of 2.08μM for peptide H2 and 8.96μM for peptide H3 were determined. Dimer formation of the C-terminal dimerization domain was analyzed by microscale thermophoresis, and a KD of 1.29nM was determined. Furthermore, microscale thermophoresis studies demonstrated a high affinity binding of H2 and H3 to the C-terminal domain, with a KD of 1.02μM and 1.46μM, respectively. ConclusionsThese results revealed the first peptidic inhibitors of Hsp90 dimerization targeting the C-terminal domain. Furthermore, it has been shown that these peptides bind to the C-terminal domain with a low micromolar affinity. General significanceThese results can be used to design and screen for small molecules that inhibit the dimerization of the C-terminal domain of Hsp90, which could open a new route for cancer therapy.

Proteomics analysis provides insight into caloric restriction mediated oxidation and expression of brain proteins associated with age-related impaired cellular processes: Mitochondrial dysfunction, glutamate dysregulation and impaired protein synthesis

Age-related impairment of functionality of the central nervous system (CNS) is associated with increased susceptibility to develop many neurodegenerative diseases. Increased oxidative stress in the CNS of aged animals is manifested by increased protein oxidation, which is believed to contribute to the age-related learning and memory deficits. Glutamate dysregulation, mitochondrial dysfunction and impaired protein synthesis are observed in aged brains, along with increased protein oxidation. Interestingly, all of these age-related cellular alterations can be improved by caloric restriction (CR), which can also improve the plasticity and recovery of the CNS. Although the beneficial effects of CR on brains are well established, the mechanism(s) of its action remains unclear. In order to gain insight into the mechanism of CR in the brain, we located the brain regions that are benefited the most from reduced oxidative stress by CR. Along with other brain regions, striatum (ST) showed significantly decreased bulk protein carbonyl levels and hippocampus (HP) showed decreased bulk protein 3-nitrotyrosine (3-NT) levels in CR aged rats when compared to those of age matched controls. To determine which proteins were oxidatively modified in these brain regions, we used parallel proteomics approach to identify the proteins that are altered in oxidation and expression. The specific carbonyl levels of pyruvate kinase M2 (PKM2), α-enolase (ENO1), inositol monophosphatase (INSP1), and F1-ATPase Chain B (ATP-F1B) were significantly decreased in ST of aged CR rats. In contrast, the expression levels of phosphoglycerate kinase 1 (PKG1), inosine monophosphate cyclohydrolase (IMPCH) and F1-ATPase Chain A (ATP-F1A) were significantly increased in the ST of CR rats. In the hippocampus of CR rats, the specific 3-NT levels of malate dehydrogenase (MDH), phosphoglycerate kinase 1 (PKG1) and 14-3-3 zeta protein were significantly decreased and expression levels of DLP1 splice variant 1 (DLP1), mitochondrial aconitase (ACO2), dihydrolipoamide dehydrogenase (DLDH), neuroprotective peptide H3 (NPH3), and eukaryotic translation initiation factor 5A (eIF-5A) are increased. Moreover, an unnamed protein product (UNP1) with similar sequence to initiation factor 2 (IF-2) was decreased in the HP of CR rats. Our data support the hypothesis that CR induces a mild metabolic stress response by increasing the production of neurotrophic proteins, therefore, priming neurons against apoptosis. Moreover, our study shows that the improvement of glutamate dysregulation, mitochondrial dysfunction and protein synthesis by CR is, at least partially, due to the CR-mediated alteration of the oxidation or the expression of PKM2, ENO1, INSP1, ATP-F1B, PKG1, IMPCH, ATP-F1A MDH, PKG1 and 14-3-3 zeta protein, DLP1, ACO2, DLDH, NPH3, eIF-5A and UNP1. This study provides valuable insights into the mechanisms of the beneficial factors on brain aging by CR.

Cytolytic and antibacterial activity of synthetic peptides derived from amoebapore, the pore-forming peptide of Entamoeba histolytica.

The pore-forming peptide amoebapore is considered part of the cytolytic armament of pathogenic Entamoeba histolytica. Amoebapore is composed of 77 amino acid residues arranged in four alpha-helical domains. For structure-function analysis, synthetic peptides were constructed corresponding to these four domains: H1 (residues 1-22), H2 (25-39), H3 (40-64), and H4 (67-77). The peptides H1 and H3, representing two highly amphipathic alpha-helical regions of amoebapore, possessed pore-forming activity. Peptide H3 displayed cytolytic and antibacterial functions similar to those of natural amoebapore. The most potent antibacterial activity and the broadest activity spectrum were expressed by H1-Mel, a hybrid molecule composed of the N-terminal alpha-helix of amoebapore and the C-terminal hexapeptide of melittin from the venom of Apis mellifera.

Minimum requirements for immunogenic and antigenic activities of homologs of a synthetic peptide of influenza virus hemagglutinin

Synthetic peptides of increasing length and corresponding in sequence to the C-terminal end of the HA1 molecule of influenza virus were constructed and examined for their immunogenic and antigenic properties. Peptides containing at least the four C-terminal amino acids, when coupled to keyhole limpet hemocyanin, were capable of eliciting antibody in BALB/c mice that bound to the 24-residue parent peptide H3 HA1 (305 to 328). In the absence of a carrier, the C-terminal decapeptide was the shortest peptide capable of eliciting antibody. The specificity of this antibody was indistinguishable from that of a monoclonal antibody to the parent peptide which recognizes an epitope encompassed by the C-terminal seven residues. All peptides containing at least the C-terminal four residues were able to inhibit completely the binding of this monoclonal antibody to the parent peptide. Taken together, these results indicate that (i) the tetrapeptide is capable of eliciting specific antibody when coupled to a carrier, (ii) this tetrapeptide possesses all of the antigenic information necessary to occupy the paratope of a monoclonal antibody elicited by the longer parent peptide, and (iii) the decapeptide contains all of the information necessary to elicit a specific immune response and therefore carries an epitope recognized by T cells as well as one recognized by B cells.

Genetic control and fine specificity of the immune response to a synthetic peptide of influenza virus hemagglutinin.

The immune response to a synthetic peptide, H3 HA1(305-328), representing the C'-terminal 24 amino acid residues of the HA1 chain of the hemagglutinin of the H3 subtype of influenza virus is controlled by genes in the I region of the major histocompatibility complex. Mice of the H-2d haplotype are high responders and produce antibody for several months after a single injection of peptide without carrier. Mice of the H-2b, H-2k, and H-2q haplotypes are low antibody responders. Investigation of recombinant and congenic mouse strains revealed that high responsiveness requires the genes that encode the I-Ed molecule. Immunoassays, involving direct binding to analogs of this peptide and inhibition by both these analogs and synthetic epitopes, were used to analyze the specificity of the polyclonal response. In BALB/c mice, the primary antibody response is directed principally against the antigenic site 314-LKLAT-318, whereas the secondary response after a boost is predominantly directed to a distinct site, 320-MRNVPEKQT-328. The T-cell response to the peptide H3 HA1(305-328), as measured by antigen-induced proliferation of primed T cells in vitro, is also I-Ed restricted in high-responder H-2d mice and is directed against an antigenic site that does not require the four C-terminal residues unique to the H3 influenza subtype. A different epitope appears to be recognized by T cells from CBA (H-2k) mice, which proliferate to a moderate extent on exposure to the peptide but, nevertheless, do not provide help for an antibody response.