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Proteinase Activity (proteinase + activity)
Selected AbstractsResponse of intestinal proteinase activities to the feeding of isolated winged bean (Psophocarpus tetragonolobus) and soya bean (Glycine max) trypsin inhibitors in ratsJOURNAL OF THE SCIENCE OF FOOD AND AGRICULTURE, Issue 11 2001Naoki Nishino Abstract The antinutritional activities of trypsin inhibitors (TIs) were compared between winged beans (Psophocarpus tetragonolobus) and soya beans (Glycine max). The inhibitors of the two beans were isolated by trypsin-bound Sepharose 4B, and 50,mg of lyophilised powders were intubated intragastrically into 24,h fasted rats. The activities of trypsin and chymotrypsin were compared after 30, 60 and 180,min in the washings of the upper, middle and lower parts of the small intestine. The elution profiles of TI and non-TI compounds in the affinity chromatography were similar in the two beans, and the antitryptic activities were concentrated 5.5 and 6.2 times (based on specific activity) for winged beans and soya beans respectively. Regardless of the TI fed to rats, trypsin activity in the upper intestine was suppressed to almost undetectable levels at 30 and 60,min after intubation. The activities in the middle and lower intestines were also substantially lowered when rats were fed winged bean TI, and significant differences were detected at 30 and 60,min after intubation when compared with rats fed soya bean TI. However, at 180,min after feeding, no differences were found in the trypsin activity in any gut segments. Similar inhibitory properties of isolated TIs were observed in chymotrypsin activities in the small intestine. The results suggest that winged bean TI may have greater inhibitory activity on the intestinal proteinase compared with soya bean TI. © 2001 Society of Chemical Industry [source] A mass spectrometry-based strategy for detecting and characterizing endogenous proteinase activities in complex biological samplesPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 3 2008Sarah Robinson Abstract Endogenous proteinases in biological fluids such as human saliva produce a rich peptide repertoire that reflects a unique combination of enzymes, substrates, and inhibitors/activators. Accordingly, this subproteome is an interesting source of biomarkers for disease processes that either directly or indirectly involve proteolysis. However, the relevant proteinases, typically very low abundance molecules, are difficult to classify and identify. We hypothesized that a sensitive technique for monitoring accumulated peptide products in an unbiased, global manner would be very useful for detecting and profiling proteolytic activities in complex biological samples. Building on the longstanding use of 18O isotope-based approaches for the classification of proteolytic and other enzymatic processes we devised a new method for evaluating endogenous proteinases. Specifically, we showed that upon ex vivo incubation endogenous proteinases in human parotid saliva introduced 18O from isotopically enriched water into the C-terminal carboxylic groups of their peptide products. Subsequent peptide sequence determination and inhibitor profiling enabled the detection of discrete subsets of proteolytic products that were generated by different enzymes. As a proof-of-principle we used one of these fingerprints to identify the relevant activity as tissue kallikrein. We termed this technique PALeO. Our results suggest that PALeO is a rapid and highly sensitive method for globally assessing proteinase activities in complex biological samples. [source] PROTEINASES IN HYBRID CATFISH VISCERA: CHARACTERIZATION AND EFFECT OF EXTRACTION MEDIAJOURNAL OF FOOD BIOCHEMISTRY, Issue 4 2010SAPPASITH KLOMKLAO ABSTRACT Proteolytic activity from viscera extract of hybrid catfish (Clarias macrocephalus × Clarias gariepinus) was investigated. Optimal pH and temperature for casein hydrolysis were 9.0 and 50C, respectively. The enzyme was stable to heat treatment up to 40C and over a pH range of 7,11 for 30,120 min. The proteolytic activity was effectively inhibited by soybean trypsin inhibitor, benzamidine, phenylmethylsulfonyl fluoride and N -p-tosyl-L-lysine chloromethyl ketone. Activities of the viscera extract continuously decreased as NaCl concentration increased, while activities increased as CaCl2 concentration increased. Based on the proteinase activity of zones separated by electrophoresis, the molecular mass of the major proteinases in hybrid catfish viscera was 23 and 20 kDa. The effect of extraction media on recovery of proteinases was also studied. Extraction of the viscera powder with 50 mM Tris-HCl, pH 7.0 containing 0.5 M NaCl and 0.2% (v/v) Brij 35 rendered a higher recovery of proteinase activity than other extractants tested (P < 0.05). The results suggested that major proteinases in hybrid catfish viscera were heat-activated alkaline proteinases, most likely trypsin-like serine proteinases. PRACTICAL APPLICATIONS Hybrid catfish viscera is an abundant and underutilized resource that can be used as a unique proteinase source. Proteinase from various sources catalyzes the hydrolysis of peptide bonds. Thus, it is expected that like other proteinases, hybrid catfish proteinase would be useful in biomedical, food and beverage application. Moreover, the presented extraction media could be adopted to recover the trypsin-like serine proteinase from hybrid catfish viscera, which is currently a solid waste of Pa-duk-ra industry. [source] Presenilin 1 mediates retinoic acid-induced differentiation of SH-SY5Y cells through facilitation of Wnt signalingJOURNAL OF NEUROSCIENCE RESEARCH, Issue 2 2003Kengo Uemura Abstract Presenilin 1 interacts with ,-catenin, an essential component of the Wnt signaling pathway. To elucidate the role of presenilin 1-,-catenin interaction in neuronal differentiation, we established SH-SY5Y cells stably expressing wild-type presenilin 1, P117L mutant presenilin 1, which is linked to the early-onset familial form of Alzheimer's disease, and D385A mutant presenilin 1, which has no aspartyl proteinase activity. We demonstrate that SH-SY5Y cells stably expressing D385A mutant presenilin 1 failed to differentiate in response to retinoic acid treatment. Retinoic acid caused an increase in nuclear ,-catenin levels in SH-SY5Y cells, which was followed by an increase in cyclin D1 protein levels. Abnormal cellular accumulation of ,-catenin was observed in D385A mutant transfected cells, whereas nuclear ,-catenin and cellular cyclin D1 levels failed to increase. Conversely, SH-SY5Y cells expressing the P117L mutant differentiated normally and showed increased nuclear ,-catenin and cellular cyclin D1 levels. These findings suggest that neuronal differentiation of SH-SY5Y cells involves the Wnt signaling pathway and that presenilin 1 plays a crucial role in Wnt signal transduction by regulating the nuclear translocation of ,-catenin. © 2003 Wiley-Liss, Inc. [source] Selective inhibition of Porphyromonas gingivalis growth by a factor Xa inhibitor, DX-9065aJOURNAL OF PERIODONTAL RESEARCH, Issue 3 2006Kenji Matsushita Background:,Porphyromonas gingivalis is a causative bacterium of adult periodontitis. However, there is no drug specific for P. gingivalis and for its virulence factor. Objectives:, The objective of this study was to examine the effects of a new selective inhibitor of activated factor X, DX-9065a, on growth of Porphyromonas gingivalis and other periodontopathic bacteria. Methods:, We incubated P. gingivalis and other periodontopathic bacteria in the presence or absence of DX-9065a and examined the effect of DX-9065a on bacterial growth and trypsin-like activity in its cultures. We also examined the effects of DX9065a on amidolytic activity of purified trypsin-like proteinases (gingipains RgpA and RgpB), from P. gingivalis and on trypsin-like activity in gingival crevicular fluids from patients with adult periodontitis. Results:, DX-9065a selectively inhibited the growth of P. gingivalis and Prevotella intermedia, and its effect on P. gingivalis was bactericidal. Trypsin-like proteinase activity was detected in P. gingivalis, and the activity was strongly inhibited by DX-9065a. DX-9065a even inhibited amidolytic activity of RgpA and RgpB from P. gingivalis. Furthermore, trypsin-like proteinase activity in gingival crevicular fluids was strongly inhibited by DX-9065a. Conclusions:, DX-9065a inhibits P. gingivalis growth in part through to its ability to inhibit the trypsin-like proteinase activity in P. gingivalis and may be useful for a new drug for treatment of adult periodontitis. [source] Changes in Wheat Leaf Extracellular Proteolytic Activity after Infection with Septoria triticiJOURNAL OF PHYTOPATHOLOGY, Issue 3 2002C. I. SEGGARRA The proteolytic activity of the leaf extracellular space of wheat cultivars Pigüé and Isla Verde was estimated after inoculation of either detached leaves or plants with the fungus Septoria tritici. Pigüé is resistant, whereas Isla Verde is susceptible to the disease caused by S. tritici. Viable conidiospores of the fungus caused similar increases in both hydrogen peroxide production and chitinase activity of the cultivars studied. In contrast, they caused a decrease in the extracellular serine proteinase activity of Isla Verde and a significant increase in that of Pigüé. Independently of the cultivar from which it was extracted, the extracellular serine proteinase inhibited the germination of Septoria tritici conidiospores. These results suggest that the proteolytic activity of the leaf extracellular space can participate in the defence of wheat plants against Septoria tritici. Its regulation may be controlled by specific defence components of each cultivar. [source] Discrepancies between the phenotypic and genotypic characterization of Lactococcus lactis cheese isolatesLETTERS IN APPLIED MICROBIOLOGY, Issue 6 2006M. De La Plaza Abstract Aims:, The use of randomly amplified polymorphic DNA (RAPD)-PCR fingerprinting and plasmid profiles to determine at the strain level, the similarity of Lactococcus lactis isolates obtained during sampling of traditional cheeses and to verify its correspondence to the selected phenotypic characteristics. Methods and Results:, A total of 45 L. lactis isolates were genotypically analysed by RAPD-PCR fingerprinting and plasmid patterns. Phenotypic traits used to compare strains were proteolytic, acidifying, aminotransferase (aromatic and branched chain aminotransferase) and , -ketoisovalerate decarboxylase (Kivd) activities. The results show that 23 isolates could be grouped in clusters that exhibited 100% identity in both their RAPD and plasmid patterns, indicating the probable isolation of dominant strains during the cheese sampling process. However, there were phenotypic differences between isolates within the same cluster that included the loss of relevant technological properties such as proteinase activity and acidifying capacity or high variation in their amino acid converting enzyme activities. Likewise, the analysis of a specific attribute, Kivd activity, indicated that 7 of 15 isolates showed no detectable activity despite the presence of the encoding (kivd) gene. Conclusion:, Phenotypic differences found between genotypically similar strains of L. lactis strains could be linked to differences in enzymatic expression. Significance and Impact of the Study:, Phenotypic analysis of L. lactis isolates should be considered when selecting strains with new cheese flavour forming capabilities. [source] Uterine secretion of ISP1 & 2 tryptases is regulated by progesterone and estrogen during pregnancy and the endometrial cycleMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 3 2004Colleen M. O'Sullivan Abstract We have described two novel implantation serine proteinase (ISP) genes that are expressed during the implantation period. The ISP1 gene may encode the embryo-derived enzyme strypsin, which is necessary for blastocyst hatching in vitro and the initiation of invasion. The ISP2 gene, which encodes a related tryptase, is expressed in endometrial glands and is regulated by progesterone during the peri-implantation period. Based on similarities between ISP2 gene expression and that of a progesterone-regulated lumenal serine proteinase activity associated with lysis of the zona pellucida, we have suggested that the strypsin related protein, ISP2, may encode a zona lysin proteinase. Recently strypsin has also been found within uterine fluid, suggesting a second potential role in hatching. Consistently, we have discovered that ISP1 is also expressed in the uterine secretory gland at the time of hatching. In this study we demonstrate that both ISP1 and ISP2 are secreted together into the uterine lumen at peri-implantation, and that the appearance of ISP protein is regulated positively at the transcriptional level by progesterone and negatively at the posttranscriptional level by estrogen. This negative regulation by estrogen may be overridden in pregnancy as ISP protein expression is restored during oil-induced decidualization. ISP1 and ISP2 proteins are also expressed in proestrous suggesting additional roles in the endometrial cycle. Mol. Reprod. Dev. 69: 252,259, 2004. © 2004 Wiley-Liss, Inc. [source] Matrix metalloproteinase-7 triggers the matricrine action of insulin-like growth factor-II via proteinase activity on insulin-like growth factor binding protein 2 in the extracellular matrixCANCER SCIENCE, Issue 5 2007Shin'ichi Miyamoto Many growth factors and cytokines are immobilized on the extracellular matrix (ECM) by binding to glycosaminoglycans and are stored in an inactive form in the cellular microenvironment. However, the mechanisms of ECM-bound growth factor or cytokine activation have not been well documented. We showed that the insulin-like growth factor type-1 receptor (IGF-1R) was rapidly phosphorylated after the addition of matrix metalloproteinase (MMP)-7 to a serum-starved human colon cancer cell line (HT29) and that phosphorylation was completely inhibited by an IGF-II neutralizing antibody. In the ECM of this cell line, IGF-II and IGF binding protein (BP)-2 coexisted, but IGFBP-2 disappeared from the ECM fraction after treatment with MMP-7 or heparinase III. On the other hand, in a cell line in which IGF-1R was overexpressed, IGF-1R was phosphorylated by supernatant from the MMP-7-treated ECM fraction of HT29 but not by that from a heparinase-III-treated ECM fraction. We also demonstrated that MMP-7 degrades IGFBP-2 in vitro at three cleavage sites (peptide bonds E151,L152, G175,L176 and K181,L182), which have not been documented previously. Taken together, these results demonstrate that MMP-7 generates bioactive IGF-II by degrading the IGF-II/IGFBP-2 complex binding to heparan sulfate proteoglycan in the ECM, resulting in IGF-II-induced signal transduction. This evidence indicates that some ECM-associated growth factors enhance their ability to bind to their receptors by some proteases in the tumor microenvironment. This mechanism of action (,protease-triggered matricrine') represents an attractive model for understanding ECM,tumor interactions. (Cancer Sci 2007; 98: 685,691) [source] Overexpression of cysteine proteinase 2 in Entamoeba histolytica or Entamoeba dispar increases amoeba-induced monolayer destruction in vitro but does not augment amoebic liver abscess formation in gerbilsCELLULAR MICROBIOLOGY, Issue 1 2001A. Hellberg To study the role of cysteine proteinases in the pathogenicity of Entamoeba histolytica, we have attempted to overexpress the three main cysteine proteinases (EhCP1, EhCP2, EhCP5) of this parasite in trophozoites of E. histolytica as well as in non-pathogenic Entamoeba dispar by episomal transfection. Although each of the corresponding coding sequences were cloned in identical expression plasmids, we were unable to overexpress EhCP1 and EhCP5, respectively, but could substantially induce expression of EhCP2 in both amoeba species by sevenfold, leading to a threefold increase in total cysteine proteinase activity. Overexpression of EhCP2 did not influence expression of other cysteine proteinases and could be attributed to an increase of a single 35 kDa activity band in substrate gel electrophoresis. In contrast to previous findings, which indicated that amoeba cysteine proteinases are involved in erythrophagocytosis and liver abscess formation, cells overexpressing EhCP2 showed no difference in erythrophagocytosis or liver abscess formation compared with respective controls. However, overexpression of EhCP2 in both amoeba species resulted in a marked increase of in vitro monolayer destruction. [source] | ||||||||||||||||