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Objective. To investigate the effects of mung bean peptide and its’ complexes on promoting lead excretion and neuroprotection of zebrafish. Methods. The lead poisoning models of zebrafish were established by lead acetate solution; the models were treated with high and low concentrations (58.3 and 175 μg/mL) of mung bean peptides, with high, medium, and low concentrations (27.8, 83.3, and 250 μg/mL) of mung bean peptide complexes, separately. The effects of the mung bean peptide complexes on the lead content, axonal fluorescence intensity, and peripheral motor nerve length changes were identified in the zebrafish model, and the effects of mung bean peptide and its’ complexes on zebrafish's lead excretion, axonal protection rate, and peripheral movement promotion rate of nerve regeneration were calculated. Results. The effects of high concentration of mung bean peptide (175 μg/mL) in promoting lead excretion was 29% ( p < 0.05 ), and the effect of high concentration of mung bean peptide complexes (250 μg/mL) in promoting lead excretion was 30% ( p < 0.05 ). The other concentrations of mung bean peptide and its’ complex groups did not show a noticeable lead excretion effect. The protective effects of mung bean peptide at concentrations of 58.3 and 175 μg/mL against zebrafish axonal injury were 98% and 101% ( p < 0.01 ), and the peripheral nerve regeneration promotion effects were 29% ( p > 0.05 ) and 42% ( p < 0.05 ), respectively. The protective effects of mung bean peptide complexes at concentrations of 27.8, 83.3, and 250 μg/mL against zebrafish axonal injury were 85%, 78%, and 93% ( p < 0.01 ); peripheral nerve regeneration promotion rates were 46%, 50%, and 50% ( p < 0.05 ). Conclusion. The mung bean peptide and its’ complexes can effectively promote the discharge of lead in the zebrafish lead poisoning and have protective and regeneration effects on zebrafish nerves.

Graphene oxide was found to be an efficient and reusable acid catalyst for the esterification reaction. A wide range of aliphatic and aromatic acids and alcohols were compatible with the standard conditions and afforded the corresponding products in good yields. The heterogeneous catalyst can be easily recovered and recycled in dichloro­ethane solvent with good catalytic activity.

A general and efficient method for the synthesis of a wide range of α-carbonyl furans is described here. The cyclization of enynone is catalyzed by AgBF4 in a fluorous biphasic system of perfluorodecalin and N,N-dimethylformamide. A variety of chain ketone and cyclic ketone substrates were used to investigate the scope of the reactions.

Allograft inflammatory factor-1 (AIF-1) plays an important role in various inflammatory conditions. Our previous study demonstrated that AIF-1 was over-expressed in the liver of BALB/c mice infected with Schistosoma japonicum and played significant role in the pathogenesis of schistosomiasis. The aim of this study was to focus on the effect of AIF-1 treatment on liver fibrosis and necrosis of BALB/c mice infected with S. japonicum. Seventy-two BALB/c mice were infected with cercariae of S. japonicum and then divided into three groups: AIF-1-treated group, saline-treated group, and control group. The vital signs, liver function, egg load, and hepatic pathological changes of the mice were assessed, and the levels of AIF-1 and TNF-α in the liver and spleen were measured at 5, 8, and 14 weeks postinfection. The treatment of AIF-1 on the mice infected with S. japonicum suppressed the expression of TNF-α and increased the effectiveness of AIF-1 in the liver and spleen at 14 weeks postinfection. Histopathological analysis and Masson trichrome staining for the liver tissues showed that the liver fibrosis and necrosis were alleviated previously compared with other infected mice at 14 weeks postinfection. The treatment of AIF-1 on the mice infected with S. japonicum can alleviate hepatic fibrosis and necrosis which indicate that AIF-1 use may prevent and cure the liver fibrosis.

Background: Daintain/AIF-1 is an inflammatory polypeptide factor/allograft inflammatory factor 1 derived from macrophages. It is characterized in APOE-/- mice as a novel inflammatory factor associated with atherosclerosis. The purpose of this study was to characterize its function in human atherosclerosis. Methods: Immunohistochemistry was used to identify the expression of daintain/AIF-1 in vessel segments within and far from atherosclerotic plaques; High-performance liquid chromatography (HPLC) was used to display the effects of daintain/AIF-1 on C-reactive protein (CRP), oxidative capacity and superoxide dismutase (SOD) in vivo; Oil Red O Staining was used to show the effects of daintain/AIF-1 on uptake of oxidized low density lipoprotein (ox-LDL) into U937 cells, a macrophage line; Western Blot was used to test scavenger receptor A (SRA) expression. Results: A high density of daintain/AIF-1 was observed in the tunica intima and media of coronary artery with atherosclerotic plaque, and fewer daintain/AIF-1 in the vessels without atherosclerotic plaque; Daintain/AIF-1 injected intravenously into BALB/c mice boosted oxidative capacity, significantly impaired SOD activities and augmented the CRP level in blood. According to the oil red O test, daintain/AIF-1 profoundly facilitated the uptake of ox-LDL in U937 macrophages and formation of foam cells in the endothelium. We also found that the molecular mechanisms are effective by promoting overexpression of SRA on macrophages. Conclusion: These findings implicate that the inflammatory factor daintain/AIF-1 is closely associated with atherogenesis, and could be further characterized as a novel risk factor for atherosclerosis

Vglycin, a novel natural polypeptide isolated from pea seeds, possesses antidiabetic properties. Our previous studies have shown that Vglycin can induce the differentiation of human colon adenocarcinoma cells. We aimed to determine the anticancer activity of Vglycin against colon cancer cells and to elucidate related apoptosis-inducing mechanisms. Treatment with purified Vglycin significantly reduced growth, viability, and colony formation of CT-26, SW480, and NCL-H716 colon cancer cells in a dose-dependent manner while down-regulating the expression of proliferating cell nuclear antigen. Mouse xenograft studies showed a 38% inhibition of colon cancer growth in mice treated with Vglycin (20 mg/kg/day) at day 21. Furthermore, the potential mechanisms involved in Vglycin-induced cell apoptosis were examined using cell cycle studies, ultrastructural examination, as well as apoptosis-associated pathway analysis. The results showed that Vglycin significantly promoted apoptosis and G1/S phase cell cycle arrest. As revealed by Western blot, the expression of CDK2 and Cyclin D1 was down-regulated in all three Vglycin-treated colon cancer cells, indicating that the CDK2/Cyclin D1 cell cycle pathway involved in the initiation and progression of colon cancer. Moreover, the inhibition of Vglycin-induced cell proliferation in colon cancer cells was accompanied by alteration of the expression levels of the apoptosis-related proteins Bax, Bcl-2 and Mcl-1, and an increase of caspase-3 activity. Together, our results suggest that Vglycin may be another plant-derived peptide that suppresses colon cancer, supporting the continued investigation of Vglycin as therapeutic agent for colon cancer. Impact statement The antidiabetic properties and the capability of inducing differentiation of human colon adenocarcinoma cells of Vglycin have been reported in our previous studies. However, the anticancer potential of Vglycin on colon cancer cells and its possible related mechanisms were still unknown. In this study, we found that Vglycin could reduce growth, viability, and colony formation or colony size of CT-26, SW480, and NCL-H716 colon cancer cells. Moreover, Vglycin decreased tumor volume by 38% in xenograft mice transplanted with CT-26 cells. The mechanisms of these phenomena may be due to the down-regulated CDK2 and Cyclin D1, G1/S phase cell cycle arrest, and the dysregulated expression of Bax, Bcl-2, and Mcl-1. The findings highlight the anticancer potential of Vglycin against colon cancer cells, and suggest Vglycin may be another colon cancer potential suppressive component of plant-derived peptides.

A large body of experimental evidence suggests that cytokines trigger pancreatic β-cell death in type 1 diabetes mellitus. Daintain/AIF-1 (Allograft Inflammatory Factor-1), a specific marker for activated macrophages, is accumulated in the pancreatic islets of pre-diabetic BB rats. In the present study, we demonstrate that daintain/AIF-1 is released into blood and the levels of daintain/AIF-1 in the blood of type 1 diabetes-prone non-obese diabetic (NOD) mice suffering from insulitis are significantly higher than that in healthy NOD mice. When injected intravenously into NOD mice, daintain/AIF-1 stimulates white blood cell proliferation, increases the concentrations of blood glucose, impairs insulin expression, up-regulates nitric oxide (NO) production in pancreases and accelerates diabetes in NOD mice, while the antibody against daintain/AIF-1 delays or prevents insulitis in NOD mice. These results imply daintain/AIF-1 triggers type 1 diabetes probably via arousing immune cells activation and induction of NO production in pancreas of NOD mice.

Pea albumin 1F (PA1F), a plant peptide isolated from pea seeds, can dramatically increase blood glucose concentration by subcutaneous injection with a dosage of 5 or 10 microg/g (body weight) in normal and type II diabetic mice (KK/upj-Ay). The voltage-dependent anion channel 1 (VDAC-1) has been identified as the PA1F binding protein from mice pancreatic cell membrane, which may be involved in the regulation of enhancing blood glucose in response to PA1F binding. The results clearly show that peptide-signaling molecules from plants can affect mammalian physiological functions, especially, in association with glucose metabolism.

Different cell line with increased thioredoxin-1 (Trx-1) showed a decreased or increased sensitivity to cell killing by cisplatin. Recently, several studies found that the subcellular localization of Trx-1 is closely associated with its functions. In this study, we explored the association of the nuclear Trx-1 with the cisplatin-mediated apoptosis of breast cancer cells MCF-7. Firstly, we found that higher total Trx-1 accompanied by no change of nuclear Trx-1 can not influence apoptosis induced by cisplatin in MCF-7 cells transferred with Trx-1 cDNA. Secondly, higher nuclear Trx-1 accompanied by no change of total Trx-1 can protect cells from apoptosis induced by cisplatin. Thirdly, high nuclear Trx-1 involves in the cisplatin-resistance in cisplatin-resistive cells. Meanwhile, we found that the mRNA level of p53 is closely correlated with the level of nuclear Trx-1. In summary, we concluded that the nuclear Trx-1 is required to resist apoptosis of MCF-7 cells induced by cisplatin, probably through up-regulating the anti-apoptotic gene, p53.

A 37 residue peptide, aglycin, has been purified from porcine intestine. The sequence is identical to that of residues 27-63 of plant albumin 1 B precursor (PA1B, chain b) from pea seeds. Aglycin resists in vitro proteolysis by pepsin, trypsin and Glu-C protease, compatible with its intestinal occurrence and an exogenous origin from plant food. When subcutaneously injected into mice (at 10 microg.g(-1) body weight), aglycin has a hyperglycemic effect resulting in a doubling of the blood glucose level within 60 min. Using surface plasmon resonance biosensor technology, an aglycin binding protein with an apparent molecular mass of 34 kDa was detected in membrane protein extracts from porcine and mice pancreas. The polypeptide was purified by affinity chromatography and identified through peptide mass fingerprinting as the voltage-dependent anion-selective channel protein 1. The results indicate that aglycin has the potential to interfere with mammalian physiology.

Replenishment of insulin-producing pancreatic β-cells would be beneficial in diabetes. The number of β-cells is maintained primarily by self-neogenesis to compensate for β-cell failure, loss or dedifferentiation. We present here a polypeptide vglycin, which was isolated and purified from germinating pea seeds. Vglycin exhibited positive effects in our diabetic models by promoting the proliferation and suppressing the apoptosis and dedifferentiation of β-cells. Vglycin promoted the restoration of β-cells in both young streptozotocin (STZ)-induced type 1 diabetic SD rats and in aged high-fat diet with (or without) STZ-induced type 2 diabetic C57BL/6 mice. We demonstrated that vglycin triggers this positive signaling by activating the insulin receptor and corresponding transcription factors. Impaired insulin sensitivity and glucose tolerance in aged T2DM mice were dramatically improved after long-term vglycin treatment, consistent with the altered level of inflammatory factor IL-1β/6. In addition, energy expenditure and body weights were significantly decreased in the mouse models after vglycin therapy. These results provide insight into the protective effects of vglycin on ameliorating β-cell function in standing glucolipotoxicity. Thus, vglycin may represent a new therapeutic agent for preventing and treating diabetes by replenishing endogenous insulin-positive cells.

PA 1b (pea albumin 1b), extracted from pea seeds, is thermostable and is multifunctional. It has an attractive peros toxicity, and is also involved in the regulation of callus growth and cell proliferation. Here we report the preparation of monoclonal antibodies (MAbs) against this peptide for further investigation of peptide distribution and functions. PA 1b was coupled to carrier protein using the two-step glutaraldehyde method as an immunal antigen. Five stable cell lines producing anti-PA 1b MAbs were obtained. We analyzed their isotypes, titer, and affinity and found that those MAbs belong to the G(1) and G(2b) subclasses with kappa light chain, respectively. Using these antibodies, a competitive inhibition ELISA was developed, and approximately 15 nmol/L of antigen was detected.

An efficient procedure for the preparation of α-acetoxy ketone derivatives by palladium-catalyzed oxidation reactions employing molecular oxygen as the sole oxidant has been reported. A variety of ...

GIP1-39, a novel chain-length form of GIP (gastric inhibitory polypeptide or glucose-dependent insulinotropic polypeptide), has been purified recently from porcine intestine and found to exist abundantly in this tissue. We have characterized that GIP1-39 is an insulinotropic peptide, and demonstrated that GIP1-39 is more potent in stimulating insulin secretion from rat pancreatic islets than GIP1-42, the insulinotropic polypeptide reported originally. Therefore, we have further investigated some aspects on the mechanism behind the insulinotropic effect of GIP1-39 in single rat pancreatic beta cells. GIP1-39 at 100 nM was able to significantly increase intracellular Ca2+ concentration ([Ca2+]i), and capable of enhancing exocytosis assessed by membrane capacitance measurement. The novel GIP1-39 might be a more optimal molecular pattern in stimulating insulin secretion and deserves to be further investigated biologically and clinically.

We investigated the effects of daintain/AIF-1, a novel inflammatory cytokine, on INS-1β cells. Cells incubated with daintain/AIF-1 showed decreased cell viability and glucose-stimulated insulin secretion, as well as upregulated apoptosis and NO production. These deleterious effects of daintain/AIF-1 indicate that daintain/AIF-1 plays important roles in the dysfunction of pancreatic β cells in type-1 diabetes.

We have isolated a posttranslationally modified form of peptide YY (PYY) from porcine intestine and shown by MALDI-TOF and electrospray tandem mass spectrometry that it is phosphorylated at Ser13. Phospho-PYY exhibits high affinity for binding to neuropeptide Y (NPY) receptors Y1, Y2 and Y5. The IC50 values with the Y1, Y2, and Y5 receptor subtypes were for NPY 2.4, 3.1, and 3.3 nM, for PYY 2.3, 0.94, and 3.2 nM, and for phospho-PYY 4.6, 2.2, and 5.5 nM, respectively. Phospho-PYY potently inhibits forskolin-stimulated cAMP accumulation in SK-N-MC cells with an IC50 value of 0.5 nM compared to 0.15 nM for non-phosphorylated PYY. The finding of phosphorylation of PYY is unusual among hormonal peptides, and emphasizes the importance of direct protein analysis of gene products.

An 84-residue bactericidal peptide, PSK, was purified from a Chrysomya megacephala fly larvae preparation. Its amino acid sequence is similar to that of a previously reported larval peptide of the Drosophila genus (SK84) noticed for its anticancer and antimicrobial properties. The PSK sequence is also homologous to mitochondrial ATPase inhibitors from insects to humans (35–65% sequence identity), indicating an intracellular protein target and possible mechanism for PSK. It contains a cluster of six glycine residues, and has several two- and three-residue repeats. It is active against both Gram-positive and Gram-negative bacteria via a mechanism apparently involving cell membrane disintegration and inhibition of ATP hydrolysis. In addition, PSK induces an inward cationic current in pancreatic β cells. Together, the findings identify a bioactive peptide of the ATPase inhibitor family with specific effects on both prokaryotic and mammalian cells.

A general and efficient method for the synthesis of a wide range of α -carbonyl furans is described here. The cyclization of enynone is catalyzed by AgBF4 in a fluorous biphasic system of perfluorodecalin and N,N-dimethylformamide. A variety of chain ketone and cyclic ketone substrates were used to investigate the scope of the reactions.

A 62-residue polypeptide, dopuin, has been isolated from pig small intestine. It is distinguished by an N-terminal part with a high content of proline (7 in a 26-residue segment), a C-terminal part with a high proportion of histidine (3 in a 9-residue segment), and six half-cystine residues in three intrachain disulphide bridges (connecting positions 22-25, 23-54 and 35-44). The Cys and Pro distributions suggest a tight and special conformation. In contrast to PEC-60 and somatostatin, it has no established inhibitory effect on insulin secretion. At 10 nM concentration, a weak inhibitory tendency is less than half of that of the other two peptides. Like gastrointestinal trefoil peptides, dopuin has three disulphide bridges, Ala-Pro segments, and many charged residues, but they are differently distributed and dopuin belongs to a separate, apparently novel family. However, dopuin is similar to a peptide corresponding to an expressed-sequence-tag cDNA of human fetal liver and spleen, establishing the nature of the mature form of the product of this cDNA, and showing a general tissue, age, and species distribution of this peptide. A truncated form of vimentin, composed of its C-terminal 37 residues, vimentin-C37, was also purified and structurally characterized. These two peptides increase the complexity of known intestinal polypeptides and at least dopuin has properties compatible with specific biofunctions.

Two forms of diazepam-binding inhibitor (DBI) have been purified from chicken intestine and identified as the intact avian polypeptide (residues 1–86) and a truncated variant (residues 35–86). At 10 nM concentration, both the intact and the truncated peptide suppress in vitro-monitored glucose-induced insulin release by 50 (p

Allograft Inflammatory Factor-1 (AIF-1) is a 17kDa cytoplasmic, calcium-binding, inflammation-responsive scaffold protein that is mainly expressed in immunocytes. AIF-1 influences the immune system at several key points and thus modulates inflammatory diseases. AIF-1 boosts the expression of inflammatory mediators such as cytokines, chemokines, inducible nitric oxide synthase (iNOS) and promotes inflammatory cell proliferation and migration. Here we provide an overview of the different pathological processes regulated by AIF-1 mainly including allograft rejection, autoimmune diseases, central nervous system (CNS) injury, vasculopathy and cancer et al.

During purification of porcine gut polypeptides with respect to glucose-induced insulin secretion from pancreatic β cells, two novel fragments of the known gastrointestinal peptides were isolated and chemically characterized. They are truncated forms of phosphatidylethanolamine-binding protein (PEBP93-124) and valosin (Valosin3-25). These novel fragments, Valosin3-25 and PEBP93-124, showed stimulatory activities on glucose-induced insulin secretion from pancreatic β cells. That the novel fragments had their respective bioactivities imply that these novel peptide-fragments could be the novel processed peptide-forms in organism. Therefore, isolation and chemical characterization of bioactive peptides from natural materials in the post genomic and bio-informational era are still necessary and will help scientist to have deeper insight into proteomics. In addition, these purified porcine gut polypeptides may provide an option to the treatment of Diabetes and also bring limelight to the mechanism of Diabetes manifestation. Keywords: Insulin, truncated peptide, phosphatidylethanolanmine binding protein, valosin, glucose-dependent insulinotropic polypeptide.

We investigated the effect of 17β-estradiol (E2) on the expression of daintain/AIF-1, a marker of activated macrophages, in RAW264.7. E2 upregulated the protein and mRNA levels of daintain/AIF-1 in similar manners under physiological concentrations of 10(-11) M to 10(-7) M. The application of ICI 182,780, an estrogen receptor (ER) antagonist, attenuated E2-induced daintain/AIF-1 production, suggesting the involvement of ER in this process.

Aims Thioredoxin-1 (Trx) is an important protein involved in the regulation of apoptosis and in enhancing drug resistance in cancer cells. Threonine100 (T100) of Trx was reported to be phosphorylated; however, the role of this phosphorylation in regulating activity remains unresolved. We explored whether and how the phosphorylation of Trx is involved in drug resistance in cancer cells. Main methods The levels of phosphorylated Trx were detected by sandwich ELISA. Cell viability was investigated using Alamar Blue assay, and apoptosis was evaluated with Hoechst33258 staining. Western blotting was used to examine the changes in the expression levels of Trx, NF-κB p65 subunit, ASK-1 and 6His tag. Additionally, we took advantage of phorbol-12-myristate-13-acetate (PMA)to activate protein kinase C (PKC) and staurosporine to inhibit PKC. Key findings Trx mutated at T100 causes lower survival rate induced by H2O2, and higher apoptosis rate induced by cis-platinum and adriamycin in HepG2 cells. T100 of Trx can be phosphorylated through the PKC-dependent pathway. Furthermore, the resistance of anticancer drugs can be decreased when the phosphorylation of T100 in Trx was blocked by staurosporine. Though the Trx-ASK1 complex is not affected, the phosphorylation contributes to the nuclear location of Trx, and then up-regulates the activity of NF-κB. Significance It was firstly found that the phosphorylation of Trx at T100 plays an important role in its cytoprotective activity in cancer cells. Thus, besides blocking the active site of Trx, inhibiting the phosphorylation of Trx at T100 may be a new potential cancer therapy target.

We report herein the isolation and characterization of a novel glycine-rich antimicrobial peptide purified from the larvae of Drosophila virilis. A range of chromatographic methods was used for isolation and its antibacterial activity against Bacillus subtilis was employed to screen for the most active fractions. The peptide, termed SK84 due to its N-terminal serine, C-terminal lysine and a total of 84 residues, was completed sequenced using RT-PCR cDNA cloning. SK84 contains a high level of glycine (15.5%) and a hexaglycine cluster motif in the N-terminal part. SK84 displayed antibacterial activity against the tested Gram-positive bacteria (B. subtilis, Bacillus thuringiensis and Staphylococcus aureus), but had no effect on Gram-negative bacteria (Pseudomonas aeruginosa, Escherichia coli) and fungi (Saccharomyces cerevisiae, Candida albicans). SK84 had specific inhibitory effects on the proliferation of several cancer cell lines (Human leukemia THP-1, liver cancer HepG2, and breast cancer MCF-7 cells), but no hemolytic activity. The results from scanning electron microscopy observations revealed that SK84 killed THP-1 cells by destroying the cell membranes. Alignment results show that SK84 is a mature protein processed from the pseudoprotein GJ19999 from D. virilis, and is very similar to several pseudoproteins from different Drosophila species. Our results show that SK84 represents a novel glycine-rich peptide family in Drosophila species with antimicrobial and anti-cancer cell activities.

SK(66)-his, a novel glycine-rich peptide derived from the CG13551 gene of Drosophila, was directly expressed in Escherichia coli with the help of the glucose effect of the lac repressor and efficiently purified in high yield (10.063 mg/l). It showed significant activity against Gram-positive bacteria. We determined a MIC value of 9 microg ml(-1) with Bacillus thuringiensis.

Recent research indicates that inflammatory factors play important roles in the initiation and progression of cancers, including breast cancer. Daintain/allograft inflammatory factor-1 (AIF-1) is a crucial mediator in the inflammatory response, but it has not yet been reported whether daintain/AIF-1 is involved in the development of breast cancers. In this study, immunohistochemical analysis found strong positive expression of daintain/AIF-1 in breast ductal tumor epithelia, but only weakly positive or negative expression in the adjacent histologically normal ductal epithelia. Then, the effect of daintian/AIF-1 on the proliferation of the breast cancer cell line MDA-MB-231 was explored via transduction of the daintian/AIF-1 gene into the cells, and via inhibition of the expression of daintain/AIF-1 through short interference RNA. The results demonstrated that up-regulation and down-regulation of daintain/AIF-1 expressions promoted and inhibited the proliferation of MDA-MB-231, respectively. More interestingly, daintain/AIF-1 overexpression facilitated tumor growth in female nude mice. Furthermore, we found that daintain/AIF-1 overexpression up-regulated the expression of cyclin D1 and enhanced the transcriptional activity of nuclear factor-kappa B (NF-kappaB), a regulator of cyclin D1 expression. In contrast, the down-regulation of daintain/AIF-1 expression decreased cyclin D1 expression and inhibited the transcriptional activity of NF-kappaB. These results strongly suggest that daintain/AIF-1 can promote the growth of breast tumors via activating NF-kappaB signaling, which consequently up-regulates the expression of cyclin D1, implying that daintain/AIF-1 may be a novel target molecule for the prognosis and therapy of breast cancer.

Thioredoxin (Trx) is a member of the thioredoxin protein family and has a conserved catalytic domain (-Trp-Cys-Gly-Pro-Cys-Lys-) with reduction/oxidation (redox) activity. There are two main members in this family, Trx-1, a cytosolic and nuclear form, and Trx -2, a mitochondrial form. Trx-1 is a 104 amino acid multifunctional protein that has been extensively studied. Here we report the preparation of monoclonal antibodies (MAb) against recombinant human Trx-1(hTrx). To enhance its immunogenicity, Trx-1 was coupled to carrier protein bovine serum albumin by a two-step glutaraldehyde method. Using conventional procedures, we prepared three stable hybridoma cell lines that can produce and secret anti-Trx MAbs. We further analyzed their isotypes, titer, and affinity and found that those MAbs belong to the G1 subclass with kappa light chains, respectively. The MAbs were capable of recognizing hTrx-1, as determined by Western blotting.

Monoclonal antibodies (MAbs) against insulin are useful for insulin assays because of their specificity and plentiful supply. We have produced four monoclonal cell strains stably secreting the monoclonal antibodies against insulin; further established subpopulation, titer, affinity; and identified the antibodies as being of subclass IgG(2b)(kappa), one strain, or IgG(1)(kappa), three strains. The smallest detectable level of human insulin by ELISA using this IgG(1) was 1.25 microg/L. The monoclonal antibodies have been used for analyzing insulin content from the sera of type 2 diabetes patients and normal subjects.

A 37 residue peptide, aglycin, has been purified from porcine intestine. The sequence is identical to that of residues 27-63 of plant albumin 1 B precursor (PA1B, chain b) from pea seeds. Aglycin resists in vitro proteolysis by pepsin, trypsin and Glu-C protease, compatible with its intestinal occurrence and an exogenous origin from plant food. When subcutaneously injected into mice (at 10 microg.g(-1) body weight), aglycin has a hyperglycemic effect resulting in a doubling of the blood glucose level within 60 min. Using surface plasmon resonance biosensor technology, an aglycin binding protein with an apparent molecular mass of 34 kDa was detected in membrane protein extracts from porcine and mice pancreas. The polypeptide was purified by affinity chromatography and identified through peptide mass fingerprinting as the voltage-dependent anion-selective channel protein 1. The results indicate that aglycin has the potential to interfere with mammalian physiology.

Using alginic acid to adsorb polypeptides at pH 2.7, we isolated a peptide pea albumin 1b (PA1b) from pea seeds. The PA1b is a single chain peptide consisting of 37 amino acid residues with 6 cysteines which constitutes the cystine-knot structure. Using microfluorometry and patch clamp techniques, we found that PA1b significantly elevated the intracellular calcium level ([Ca2+]i) and elicited membrane capacitance increase in the primary rat pancreatic beta cells. The PA1b effect on [Ca2+]i elevation was abolished in the absence of extracellular Ca2+ or in the presence of L-type Ca2+ channel blocker, nimodipine. Interestingly, we found that PA1b significantly depolarized membrane potential, which could lead to the opening of voltage-dependent L-type Ca2+ channels and influx of extracellular Ca2+, and then evoke robust secretion. In this study we identified the plant peptide PA1b which is capable of affecting the excitability and function of mammalian pancreatic beta cell.

Musca domestica,which belongs to insecta, diptera, cyclorrhapha, muscidae, is the most common muscae and the richest resource. It is very significant and valuable to isolate antibacterial peptides from Musca domestica and to develop these peptides into antibacterial medicine. Due to purify a pure peptide from the natural materials (animal, plant and microorganism tissue) is very difficult and complex, few research is going on. It had been reported that the most antibacterial peptides from Musca domestica were alkaline, no weak-acid antibacterial peptides had been reported so far. Based on a high sensitivity detection method, using dilute acetic acid extraction, alginic acid absorption, NaCl salting-out, Sephadex G-25 gel filtration, CMC23 ion-exchange chromatography, electrophoresis, a group of weak-acid antibacterial peptides had been purified from Musca domestica larvae and partial characterized. The peptides had characters of broad antibacterial spectrum and low minimum bactericidal concentration against Gram-positive bacterium such as Bacillus thuringiensis, Bacillus subtilis, Staphylococcus aureus and Gram-negative bacterium such as Pseudomonas aeruginosa, Escherichia coli. The peptides were very stable to keep the antibacterial activity even kept in 95 degrees C for 120 min and frozen-thawed for 10 times. A weak-acid antibacterial peptide MD7095 had been purified in high degree of purity by electro-elution,and was determined Mr 7095Da with MALDI-TOF-MS and pl 5.59 with IEF-PAGE. Peptide mass fingerprinting (PMF) analysis showed MD7095 was a novel bioactive peptide. Few peptides with antibacterial activity against Bacillus thuringiensis had been reported. Observation by scanning electron microscopy (SEM), it was suggested that the bioactivity mechanism of antibacterial peptides from Musca domestica larvae against Bacillus thuringiensis was to perforate cell membrane and lead to bacterium lysis and die. It is hopeful to develop the antibacterial peptides from Musca domestica to candidate medicine.

A bioactive macrophage factor, the polypeptide daintain/allograft inflammatory factor 1 (AIF1), has been isolated from porcine intestine. It was discovered when searching for intestinal peptides with effects on insulin release, and its purification was monitored by the influence of the peptide fractions on pancreatic glucose-induced insulin secretion. Daintain/AIF1 is a 146-aa residue polypeptide with a mass of 16,603 Da and an acetylated N terminus. An internal 44-residue segment with the sequence pattern –KR–KK–GKR– has a motif typical of peptide hormone precursors, i.e., dibasic sites for potential activation cleavages and at the sequentially last such site, the structure GKR. The latter is a signal for C-terminal amide formation in the processing of peptide hormones. Daintain/AIF1 is immunohistochemically localized to microglial cells in the central nervous system and to dendritic cells and macrophages in several organs. A particularly dense accumulation of daintain/AIF1-immunoreactive macrophages was observed in the insulitis affecting the pancreatic islets of prediabetic BB rats. When injected intravenously in mice, daintain/AIF1 at 75 pmol/kg inhibited glucose (1 g/kg)-stimulated insulin secretion, with a concomitant impairment of the glucose elimination, whereas at higher doses (7.5 and 75 nmol/kg), daintain/AIF1 potentiated glucose-stimulated insulin secretion and enhanced the glucose elimination. Its dual influence on insulin secretion in vivo at different peptide concentrations, and the abundance of macrophages expressing daintain/AIF1 in the pancreatic islets of prediabetic rats, suggest that daintain/AIF1 may have a role in connection with the pathogenesis of insulin-dependent diabetes mellitus.

The title compound, C18H14O2, is almost planar with a dihedral angle of 1.24 (2)° between the phenylethynyl and styryl groups. The acetoxy group is tilted by 82.46 (2) and 82.26 (3)° with respect to the benzene ring planes.

Free heme is potentially cytotoxic, particularly in the presence of oxidants or activated phagocytes. Daintain/AIF-1 (allograft inflammatory factor-1) is a macrophage factor that has been implicated in the regulation of inflammation. In the present study, daintain/AIF-1 was found to induce cytolysis of erythrocytes, resulting in heme release in vitro. Furthermore, the interacting protein of daintain/AIF-1 was purified by daintain/AIF-1-6 histidine antigen fusion protein nickel affinity chromatography. MALDI-TOF-MS analysis identified hemoglobin subunit β-1 as an interacting protein of daintain/AIF-1. These data suggest that daintain/AIF-1 may be involved in heme-associated diseases