Bowman-Birk Soybean Trypsin Inhibitor Research Articles

Chymase is a potent chemoattractant for human monocytes and neutrophils.

Chymase is a major chymotrypsin-like serine protease expressed in the secretory granules of mast cells in many mammalian species. In this study, we revealed the chemotactic activity of chymase for human mononuclear cells and neutrophils with a 48-well microchemotaxis chamber technique. Human chymase showed the potent chemotactic activity for monocytes and neutrophils dose-dependently in a concentration range from 0.1 to 10 microg/mL, corresponding to about 4-400 microM. The activity was as potent as that of N-formyl-methionyl-leucyl-phenylalanine. Chymase also stimulated cell migration of lymphocytes and purified T cells, but checkerboard analysis revealed that the effect was chemokinetic rather than chemotactic. Inhibition of chymase activities with chymase inhibitors, such as antileukoprotease and Bowman-Birk soybean trypsin inhibitor, significantly inhibited the chemotactic activity of chymase, suggesting that the proteolytic activity of chymase participates in the chemotactic activity. Our results suggest that mast cell chymase acts as a chemoattractant, and may play a role in the accumulation of inflammatory cells in development of the chronic inflammatory responses of allergic and nonallergic diseases.

Purification and characterization of a novel isoform of mast cell tryptase from rat tongue.

Rat mast cell tryptase was purified to homogeneity from rat tongue by a series of standard chromatographic procedures. Since the enzyme gave band corresponding to molecular mass of 32-35 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis and exhibited a molecular mass of 135 kDa on gel filtration, it was presumed to be a noncovalently associated tetramer. The N-terminal amino acid sequence of 50 residues of the enzyme showed the highest degree of homology with the same region in mouse mast cell protease 7 (92%), and less homology to those of tryptases from man and dog, and peritoneal cells of rats and Mongolian gerbils. The inhibitor specificity of rat tongue tryptase was similar to that of rat peritoneal mast cell tryptase free from trypstatin: it was inhibited by alpha 1-antitrypsin, Kunitz-type soybean trypsin inhibitor and Bowman-Birk soybean trypsin inhibitor, but these inhibitors do not inhibit the tryptases from rat skin, human lung, and dog mast cells. Judging from these results, together with other enzymatic properties, the enzyme may be a novel isoform of tryptase in rat tongue. Analysis by differential staining with peroxidase-labeled lectins of the enzyme suggested that it has tri- and/or tetraantennary complex-type oligosaccharides containing a relatively high amount of sialic acid. The immunohistochemical distribution of this enzyme indicated that the reactive antigen was specific in connective tissue but not in mucosal mast cells.

Isolation and inhibition of a trypsin-like activity from larvae of corn borer (Ostrinia nubilalis) using reverse micelles

Endoproteinase(s) was isolated from a freeze-dried powder of larvae of Ostrinia nubilalis using reverse micellar solutions. The inhibition of proteinase was studied in reverse micelles with commercial Bowman-Birk soybean trypsin inhibitor and three trypsin inhibitors recently isolated from ripe cruciferous seeds.

New agents for cancer chemoprevention.

Clinical chemoprevention trials of more than 30 agents and agent combinations are now in progress or being planned. The most advanced agents are well known and are in large Phase III chemoprevention intervention trials or epidemiological studies. These drugs include several retinoids [e.g., retinol, retinyl palmitate, all-trans-retinoic acid, and 13-cis-retinoic acid], calcium, Beta carotene, vitamin E, tamoxifen, and finasteride. Other newer agents are currently being evaluated in or being considered for Phase II and early Phase III chemoprevention trials. Prominent in this group are all-trans-N-(4-hydroxy phenyl)retinamide (4-HPR) (alone and in combination with tamoxifen), 2-difluoromethylornithine (DFMO), nonsteroidal antiinflammatory drugs (aspirin, piroxicam, sulindac), oltipraz, and dehydroepiandrostenedione (DHEA). A third group is new agents showing chemopreventive activity in animal models, epidemiological studies, or in pilot clinical intervention studies. They are now in preclinical toxicology testing or Phase I safety and pharmacokinetics trials preparatory to chemoprevention efficacy trials. These agents include S-allyl-l-cysteine, curcumin, DHEA analog 8354 (fluasterone), genistein, ibuprofen, indole-3-carbinol, perillyl alcohol, phenethyl isothiocyanate, 9-cis-retinoic acid, sulindac sulfone, tea extracts, ursodiol, vitamin D analogs, and p-xylyl selenocyanate. A new generation of agents and agent combinations will soon enter clinical chemoprevention studies based primarily on promising chemopreventive activity in animal models and in mechanistic studies. Among these agents are more efficacious analogs of known chemopreventive drugs including novel carotenoids (e.g., alpha-carotene and lutein). Also included are safer analogs which retain the chemopreventive efficacy of the parent drug such as vitamin D3 analogs. Other agents of high interest are aromatase inhibitors (e.g., (+)-vorozole), and protease inhibitors (e.g., Bowman-Birk soybean trypsin inhibitor). Combinations are also being considered, such as vitamin E with l-selenomethionine. Analysis of signal transduction pathways is beginning to yield classes of potentially active and selective chemopreventive drugs. Examples are ras isoprenylation and epidermal growth factor receptor inhibitors.

In vivo and in vitro effects of wheat germ agglutinin and Bowman-Birk soybean trypsin inhibitor, two potential transgene products, on midgut esterase and protease activities from Apis mellifera

Wheat germ agglutinin (WGA) and Bowman-Birk soybean trypsin inhibitor represent potential transgene products for inducing pest resistance in plants. The effects of these molecules were studied on midgut esterase and protease activities from Apis mellifera L., a major insect pollinator. Trypsin inhibitor and WGA did not exhibit an acute toxicity in A. mellifera. In vivo, trypsin inhibitor caused a decrease in the amount of trypsin activity and did not have a significant effect on esterase activity. In vitro, trypsin inhibitor inhibited about 80% of non-specific protease activity and 100% of trypsin activity. In vivo, WGA at high concentration in food (1 mg/ml) elicited a large decrease in trypsin activity and did not have a significant effect on esterase activity. In vitro, WGA did not have any significant effect on trypsin and non-specific protease activities but slightly activated esterase activity.

Effect of thioredoxin-linked reduction on the activity and stability of the Kunitz and Bowman-Birk soybean trypsin inhibitor proteins

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTEffect of thioredoxin-linked reduction on the activity and stability of the Kunitz and Bowman-Birk soybean trypsin inhibitor proteinsJin An. Jiao, Boihon C. Yee, Karoly. Kobrehel, and Bob B. BuchananCite this: J. Agric. Food Chem. 1992, 40, 12, 2333–2336Publication Date (Print):December 1, 1992Publication History Published online1 May 2002Published inissue 1 December 1992https://doi.org/10.1021/jf00024a002RIGHTS & PERMISSIONSArticle Views243Altmetric-Citations42LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (503 KB) Get e-Alerts Get e-Alerts

Isolation and characterization of a novel trypsin-like protease found in rat bronchiolar epithelial Clara cells. A possible activator of the viral fusion glycoprotein.

A novel trypsin-like protease associated with rat bronchiolar epithelial Clara cells, named Tryptase Clara, was purified to homogeneity from rat lung by a series of standard chromatographic procedures. The enzyme has apparent molecular masses of 180 +/- 16 kDa on gel filtration and 30 +/- 1.5 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. Its isoelectric point is pH 4.75. Studies with model peptide substrates showed that the enzyme preferentially recognizes a single arginine cleavage site, cleaving Boc-Gln-Ala-Arg-4-methylcoumaryl-7-amide most efficiently and having a pH optimum of 7.5 with this substrate. The enzyme is strongly inhibited by aprotinin, diisopropylfluorophosphate, antipain, leupeptin, and Kunitz-type soybean trypsin inhibitor, but inhibited only slightly by Bowman-Birk soybean trypsin inhibitor, benzamidine, and alpha 1-antitrypsin. Immunohistochemical studies indicated that the enzyme is located exclusively in the bronchiolar epithelial Clara cells and colocalized with surfactant. An immunoreactive protein with a molecular mass of 28.5 kDa was also detected in airway secretions by Western blotting analyses, suggesting that the 30-kDa protease in Clara cells is processed before or after its secretion. Proteolytic cleavage of the hemagglutinin of influenza virus is a prerequisite for the virus to become infectious. Tryptase Clara was shown to cleave the hemagglutinin and activate infectivity of influenza A virus in a dose-dependent way. These results suggest that the enzyme is a possible activator of inactive viral fusion glycoprotein in the respiratory tract and thus responsible for pneumopathogenicity of the virus.

Initiation of the Degradation of the Soybean Kunitz and Bowman-Birk Trypsin Inhibitors by a Cysteine Protease

Protease K1 activity initiates the degradation of the Kunitz soybean trypsin inhibitor (KSTI) during germination and early seedling growth. This enzyme was purified nearly 1300-fold from the cotyledons of 4-day-old soybean (Glycine max [L.] Merrill) seedlings. Protease K1 is a cysteine protease with a molecular weight of approximately 29,000. It cleaves the native form of KSTI, Ti(a), to Ti(a) (m), the same modified form observed in vivo. In addition to attacking KSTI, protease K1 is also active toward the major Bowman-Birk soybean trypsin inhibitor, as well as the alpha, alpha', and beta subunits of soybean beta-conglycinin. The properties and temporal variation of protease K1 during germination indicate that it is responsible for initiating the degradation of both KSTI and Bowman-Birk soybean trypsin inhibitor in the soybean cotyledon.

Degradation of trypsin inhibitors during soybean germination

The principal Bowman-Birk soybean trypsin inhibitor in the soybean seed, BBSTI-E, is converted to BBSTI-D by limited proteolytic removal of the last two amino acid residues at the carboxyl terminus. The enzyme that catalyses this activity, Protease B1, works at an optimum pH of 4, first appears at day one and peaks at day four. The data also suggest that a second enzyme, Protease B2, appears by day 9 and catalyses extensive proteolysis of both BBSTI-E and BBSTI-D, the latter at a much faster rate. Three separate activities, measured at pH 7 in the presence of sulphydryl reducing agents, with peaks at days one, three and eight, catalyse extensive proteolysis of BBSTI-D, BBSTI-E and of BBSTI-E and -D, respectively. Immunostained Western blots show that other Bowman-Birk isoinhibitors are also digested during germination and early growth.

Proteolysis of soybean bowman-birk trypsin inhibitor during germination

The Bowman—Birk type trypsin inhibitor, BBSTI-D, which appears in the cotyledons of germinated soybeans ( Glycine max), was isolated in homogeneous form. BBSTI-D has an amino acid composition identical to the native Bowman—Birk soybean trypsin inhibitor (BBSTI-E) except for the loss of one glutamyl/glutaminyl residue and one aspartyl/asparaginyl residue. The amino-terminal sequence of BBSTI-D was identical to that of BBSTI-E. These data, as well as the compositions of the tryptic peptides from reduced carboxymethylated BBSTI-D, indicate that BBSTI-D is derived from BBSTI-E by the loss of the carboxyl-terminal residues Glu 70—Asn 71.

Differential Expression of Kunitz and Bowman-Birk Soybean Proteinase Inhibitors in Plant and Callus Tissue

Bowman-Birk soybean trypsin inhibitor (BBSTI) but not Kunitz soybean trypsin inhibitor (KSTI) was found in samples of undifferentiated and partially differentiated Amsoy 71 tissue culture callus. This suggests the differential metabolism of these two classes of proteinase inhibitors, whether the difference be in synthesis, in rates of degradation, or both. The differential metabolism of the proteinase inhibitors is also seen in the plant. Both BBSTI and KSTI were found in the hypocotyl, root, and epicotyl of the Amsoy 71 soybean seedling in addition to their expected presence in the cotyledons. Whereas the ratio of KSTI to BBSTI in the cotyledon was higher, the ratio of BBSTI to KSTI was higher in the extracotyledonary tissues of the seedling. The levels of both classes of proteinase inhibitors declined during seedling growth, except in the epicotyl and the proximal root. In both of these tissues, an increase in BBSTI, but not in KSTI content, expressed as milligrams inhibitor per plant part, occurred.

The Use of Soybean Trypsin Inhibitors as Phosphorylatable Substrates for a Rat Liver Protein Kinase

Phosphorylation by a cAMP-independent rat liver protein kinase of protein substrates containing the structural feature required by mammary gland casein kinase (-Ser-X-Glu/Asp) has been demonstrated. In particular, the Bowman-Birk Soybean trypsin inhibitor, which is characterized, like other legume protease inhibitors, by clusters of acidic residues near the C-terminal side of seryl residue(s), proved to be a good model substrate for the protein kinase. Its phosphorylation, involving the Ser 65 residue, is apparently hindered by the binding of trypsin, while it is stimulated by unfolding induced by reduction and subsequent carboxy-methylation.

Origins of circular dichroism bands in Bowman-Birk soybean trypsin inhibitor.

The spectral properties of Bowman-Birk soybean trypsin inhibitor (BBI) were investigated by analyzing difference absorption spectra and difference CD spectra and by comparing them with those of tyrosyl model compounds. The O-acetylation of tyrosyl side chains showed that the ultraviolet CD bands of BBI above 225 nm originate from disulfide side chains and tyrosyl phenolic groups; phenylalanyl residues do not give rise to detectable CD in BBI in this wavelength region. The results of the tyrosyl ionization experiment were consistent with this interpretation. A broad negative CD band centered around 280 nm in BBI arises mainly from disulfide bonds (epsilonL - epsilonR = -0.83 M-1 cm-1 per disulfide). Each of 2 tyrosyl residues gives rise to negative CD in this region; together they contribute approximately 10% of the total CD intensity at 277 nm (epsilonL - epsilonR = -0.36 M-1 cm-1 per tyrosyl). Disulfide bonds in BBI also have a broad positive CD band centered around 240 nm (epsilonL- epsilonR = 0.9 M-1 per disulfide(. Tyrosyl side chains give rise to a sharp positive peak at 231 nm, overlapping with the positive disulfide CD. Dimerization of monomeric BBI did not alter the CD profile. One of two tyrosyl phenolic groups is relatively exposed and can be 0-acetylated by 100- to 1500-fold molar excess of N-acetylmidazole. The other is inaccessible to the reagent even in the presence of 8 M urea, but can be acetylated in the presence of 6 M guanidine hydrochloride. Fully acetylated BBI has the near-ultraviolet disulfide CD and the far-ultraviolet polypetide CD very similar to those of the native inhibitor, indicating the O-acetylation of two tryosyl side chains did not induce much conformational change in BBI. The near-ultraviolet CD of BBI was altered in the presence of 8 M urea of 6 M guanidine hydrochloride, with a greater change brought about by the latter. Dithiothreitol (20 mM) completely abolished the tyrosyl and disulfide CD in this region.