Two new isomeric modified tripeptides, aspergillamides C and D (compounds 1 and 2), together with fifteen known compounds (compounds 3-17), were obtained from the marine sponge-derived fungus Aspergillus terreus SCSIO 41008. The structures of the new compounds, including absolute configurations, were determined by extensive analyses of spectroscopic data (NMR, MS, UV, and IR) and comparisons between the calculated and experimental electronic circular dichroism (ECD) spectra. Butyrolactone I (compound 11) exhibited strong inhibitory effects against Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) with the IC being 5.11 ± 0.53 μmol·L, and acted as a noncompetitive inhibitor based on kinetic analysis.
4-Butyrolactone ; analogs & derivatives ; chemistry ; isolation & purification ; pharmacology ; Animals ; Aspergillus ; chemistry ; Chemistry Techniques, Analytical ; Dipeptides ; chemistry ; isolation & purification ; pharmacology ; Enzyme Inhibitors ; chemistry ; isolation & purification ; pharmacology ; Indoles ; chemistry ; isolation & purification ; pharmacology ; Molecular Structure ; Mycobacterium tuberculosis ; drug effects ; Peptides ; chemistry ; isolation & purification ; pharmacology ; Polyketides ; chemistry ; isolation & purification ; pharmacology ; Porifera ; microbiology ; Protein Tyrosine Phosphatases ; chemistry

By using an RAD peptide display system derived from the ATPase domain of recombinase RadA of Pyrococcus furiosus, an anti-hCG antibody-like molecule was prepared by grafting an hCG-binding peptide to the RAD scaffold. After linking to sfGFP gene, a gene of hCG peptide-grafted RAD was synthesized and cloned into a bacterial expression vector (pET30a-RAD/hCGBP-sfGFP). The vector was transformed into Escherichia coli, and expression of the fusion protein was induced. After isolation and purification of the fusion protein, its binding affinity and specificity to hCG were determined by using a process of immunoabsorption followed by GFP fluorescence measurement. A comparison of hCG-binding activity with a similarly grafted single-domain antibody based on a universal scaffold was performed. The measurement of hCG-binding affinity and specificity revealed that the grafted RAD has an optimally high binding affinity and specificity to hCG, which are better than the grafted single-domain antibody. Moreover, the affinity and specificity of grafted RAD molecule are comparable to those of a commercial monoclonal antibody. In addition, the hCG-binding peptide-grafted RAD molecule has a relatively high biochemical stability, making it a good substitute for antibody with potential application.
Antibodies, Monoclonal ; chemistry ; isolation & purification ; metabolism ; Antibody Specificity ; DNA-Binding Proteins ; genetics ; metabolism ; Escherichia coli ; genetics ; Escherichia coli Proteins ; metabolism ; Humans ; Peptides ; Recombinant Fusion Proteins ; genetics ; metabolism

Two cyclopeptides, celogentin L (1) and its epimer lyciumin A (2) were firstly isolated from Celosia argentea L.. The planar structures of the two compounds were fully determined by spectroscopic data, including 1D-, 2D-NMR, and HR-ESI/MS. The absolute configurations of amino acid components were assigned via chiral-phase HPLC analyses after acid hydrolysis. Furthermore, the configuration of C-N linkage at the glycine Cα was elucidated by extensive analyses of 2D-NMR and comparison of the experimental and calculated electronic circular dichroism (ECD) spectra. Cytotoxicity of the two compounds against human alveolar epithelial A549, hepatocellular carcinoma HepG2, and cervical cancer Hela cell lines was assayed. Although both of them were inactive in these cells, the present findings add new facets for the chemistry of Celosia argentea.
A549 Cells ; Cell Survival ; drug effects ; Celosia ; chemistry ; Chemistry Techniques, Analytical ; HeLa Cells ; Hep G2 Cells ; Humans ; Molecular Conformation ; Molecular Structure ; Peptides, Cyclic ; chemistry ; isolation & purification ; toxicity ; Seeds ; chemistry ; Stereoisomerism

Herbal extracts have been extensively used worldwide for their application on memory improvement, especially among aged and memory-deficit populations. In the present study, the memory loss induced by human Abeta protein over-expression in fruitfly Alzheimer's disease (AD) model was rescued by multiple extracts from Gardenia jasminoides. Three extracts that rich with gardenia yellow, geniposide, and gardenoside components showed distinct rescue effect on memory loss. Further investigation on adding gardenoside into a formula of Ganoderma lucidum, Panax notoginseng and Panax ginseng (GPP) also support its therapeutic effects on memory improvement. Interestingly, the application of GPP and gardenoside did not alter the accumulation of Abeta proteins but suppressed the expression of immune-related genes in the brain. These results revealed the importance and relevancy of anti-inflammation process and the underlying mechanisms on rescuing memory deficits, suggesting the potential therapeutic use of the improved GPP formulation in improving cognition in defined population in the future.
Alzheimer Disease ; drug therapy ; Animals ; Antimicrobial Cationic Peptides ; genetics ; Brain ; drug effects ; immunology ; Cognition ; drug effects ; Disease Models, Animal ; Drosophila ; Drosophila Proteins ; genetics ; Gardenia ; chemistry ; Gene Expression Regulation ; drug effects ; Immunity, Innate ; drug effects ; Iridoids ; chemistry ; isolation & purification ; pharmacology ; Plant Extracts ; chemistry ; isolation & purification ; pharmacology ; Polymerase Chain Reaction

Amphibian skin contains rich bioactive peptides. Especially, a large amount of antimicrobial peptides have been identified from amphibian skin secretions. Antimicrobial peptides display potent cytolytic activities against a range of pathogenic bacteria and fungi and play important defense roles. No antimicrobial peptides have been reported from toads belonging to the family of Pelobatidae. In this work, two novel antimicrobial peptides (Megin 1 and Megin 2) were purified and characterized from the skin venoms of spadefoot toad Megophrys minor (Pelobatidae, Anura, Amphibia). Megin 1 had an amino acid sequence of FLKGCWTKWYSLKPKCPF-NH2, which was composed of 18 amino acid residues and contained an intra-molecular disulfide bridge and an amidated C-terminus. Megin 2 had an amino acid sequence of FFVLKFLLKWAGKVGLEHLACKFKNWC, which was composed of 27 amino acid residues and contained an intra-molecular disulfide bridge. Both Megin 1 and Megin 2 showed potential antimicrobial abilities against bacteria and fungi. The MICs of Megin 1 against Escherichia coli, Bacillus dysenteriae, Staphylococcus aureus, Bacillus subtilis, and Candida albicans were 25, 3, 6.25, 3, and 50 μg·mL(-1), respectively. The corresponding MICs for Megin 2 were 6.25, 1.5, 12.5, 1.5, and 12.5 μg·mL(-1), respectively. They also exerted strong hemolytic activity against human and rabbit red cells. The results suggested that megin peptides in the toad skin of M. minor displayed toxic effects on both eukaryotes and prokaryotes. This was the first report of antimicrobial peptides from amphibians belonging to the family of Pelobatidae.
Amino Acid Sequence ; Amphibian Venoms ; chemistry ; immunology ; isolation & purification ; Animals ; Anura ; immunology ; Bacillus ; Candida albicans ; Erythrocytes ; physiology ; Escherichia coli ; Female ; Hemolysis ; Humans ; Male ; Peptides ; chemistry ; immunology ; isolation & purification ; Rabbits ; Sequence Alignment ; Skin ; chemistry ; immunology ; Staphylococcus aureus

To develop a timesaving and easy operating Reverse Phase (RP) chromatography method, we adopted Thermo Pierce RP C18 Tip to separate small amount hippocampus peptide mixtures and to compare with high performance liquid chromatography (HPLC). According to the separation performance of 4 ACN gradient optimization methods, we determined the best ACN concentration gradient. The results showed that, the experiment took only 10 min by separating with eight ACN concentration gradient, which accounted 1/4 for HPLC. But as for the identified proteins, RP C18 Tip accounted 85.5% for HPLC. ACN gradient of 5%, 15%, 20% and 90% had best repeatability (P = 0.429) and result for separating 30 μg peptides. This method is easy to operate, timesaving and has low cost. It could be used into pretreatment of small amount complex proteomic samples.
Chromatography, High Pressure Liquid ; Chromatography, Reverse-Phase ; methods ; Peptides ; Proteins ; isolation & purification ; Proteomics ; methods

Enramycin is a polypeptide antibiotic and new, safe animal feed additive. A new purification process was developed, based on pre-purification by macroporous resin and refining by reversed phase chromatography. AB-8 macroporous resin was used for the pre-purification process of enramycin, with an elution buffer of 0.012 mol/L aqueous HCl solution-methanol (50: 50, V/V). Then, enramycin a and enramycin b were separated effectively by C18 reversed phase chromatography, with a elution buffer of 0.05 mol/L aqueous KH2PO4 solution-acetonitrile (70: 30, V/V, pH 4.5). The purities of enramycin a and enramycin b were up to 98.5% and 98.0%, respectively. The yield reached 29.2%. This study would provide a useful reference for the preparation of enramycin a and enramycin b with a high purity.
Adsorption ; Anti-Bacterial Agents ; isolation & purification ; Chromatography, Reverse-Phase ; methods ; Peptides ; isolation & purification

Here we discuss whether N terminal sequencing is appropriate as one of the conventional control methods for monoclonal antibody products. We determined the N terminal sequences of two monoclonal antibody products targeting two antigens separately with both Edman degradation and mass peptide spectrometry. We also identified the characteristic peptide fragments with mass spectrometry. Furthermore, we analyzed their heterogeneity with ion exchange chromatography, capillary zone electrophoresis and Imaged Capillary Isoelectric Focusing. Edman degradation method showed that the N terminal 15 amino acids of heavy and light chains of the two monoclonal antibodies were identical. Peptide mass spectrometry demonstrated that T1 peptide fragments of heavy and light chains of the two antibodies were also the same. But in contrast, peptide mapping and the three analytical methods for heterogeneity analysis could effectively identify and differentiate the two antibodies. The N terminal sequences of two monoclonal antibodies are identical because the number of framework sequences of humanized or human monoclonal antibodies is relatively limited, so whether N terminal sequencing analysis could be regulated as one of the practical control methods should be carefully discussed. Our work also proves that the above analytical methods could combinatorially applied to the identification of monoclonal antibody products, and are more objective compared to N terminal sequencing.
Amino Acid Sequence ; Antibodies, Monoclonal ; isolation & purification ; Chromatography, Ion Exchange ; Humans ; Isoelectric Focusing ; Mass Spectrometry ; Peptide Mapping ; Peptides ; Sequence Analysis, Protein ; methods

Antimicrobial Cationic Peptides ; metabolism ; Fermentation ; Food Microbiology ; India ; Lactobacillales ; isolation & purification ; metabolism ; Microbial Sensitivity Tests ; Vegetables ; microbiology

Antimicrobial Cationic Peptides ; biosynthesis ; genetics ; DNA-Binding Proteins ; biosynthesis ; genetics ; Humans ; Microbial Sensitivity Tests ; Muramidase ; biosynthesis ; genetics ; Peptides, Cyclic ; biosynthesis ; genetics ; Pichia ; metabolism ; Recombinant Proteins ; biosynthesis ; genetics ; isolation & purification