title: In vitro inhibition of the activation of Pro-matrix Metalloproteinase 1 (Pro-MMP-1) and Pro-matrix metalloproteinase 9 (Pro-MMP-9) by rice and soybean Bowman-Birk inhibitors.
abstract: The in vitro inhibitory activity of the rice Bowman-Birk inhibitor (rBBI) or soybean Bowman-Birk inhibitor (sBBI) against trypsin-catalyzed activation of pro-matrix metalloproteinase 1 or 9 (pro-MMP-1 or pro-MMP-9), respectively, was investigated using electrophoresis with silver staining, heparin-enhanced zymography, biotinylated gelatin, Biotrak assay, and fluorescence quenched substrate hydrolysis. rBBI at concentrations of 0.08-0.352 mg/mL dose-dependently inhibited the in vitro activation of 45 microg/mL pro-MMP-1 by trypsin. Heparin-enhanced zymography analysis of pro-MMP-1, trypsin-activated MMP-1, and a mixture of pro-MMP-1-trypsin-rBBI showed clear zones associated with trypsin-activated MMP-1 and the absence of clear zones in lanes containing pro-MMP-1 or a mixture of pro-MMP-1, trypsin, and rBBI. The results of the Biotrak assay also indicated that rBBI dose-dependently suppressed the activation of pro-MMP-1 by trypsin. sBBI dose-dependently inhibited the activation of 100 microg/mL of pro-MMP-9 by trypsin. Biotinylated gelatin assays demonstrated that pro-MMP-9 or pro-MMP-9 in the presence of trypsin and BBI did not hydrolyze gelatin, whereas p-aminophenylmercury acetate (APMA)-activated MMP-9 and trypsin-activated MMP-9 caused significant hydrolysis of gelatin. Quenched fluorescence substrate hydrolysis for total MMP activity showed that pro-MMP-1 or pro-MMP-9 did not hydrolyze the substrate Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2; active MMP-1 or MMP-9 hydrolyzed the substrate, but lower substrate hydrolysis was obtained when pro-MMP-1 or pro-MMP-9 was incubated with trypsin in the presence of increasing concentrations of rBBI. The results are discussed in light of the role of MMP-1 and MMP-9 in the process of angiogenesis and the potential of rBBI or sBBI as a functional food ingredient.
authors.affiliation: School of Human Ecology and Department of Food Science, Louisiana State University Agricultural Center, Baton Rouge, Louisiana 70803, USA.
authors.forename: Hiba A
authors.initials: HA
authors.lastname: Bawadi
authors.forename: Tania Maria
authors.initials: TM
authors.lastname: Antunes
authors.forename: Fred
authors.initials: F
authors.lastname: Shih
authors.forename: Jack N
authors.initials: JN
authors.lastname: Losso
mesh_terms: D019921:Biotinylation; D017364:Collagenases / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000037:antagonists & inhibitors* / Q000378:metabolism; D005453:Fluorescence; D020781:Matrix Metalloproteinase 1; D020780:Matrix Metalloproteinase 9; D061965:Matrix Metalloproteinase Inhibitors; D012275:Oryza / Q000737:chemistry*; D011480:Protease Inhibitors / Q000494:pharmacology*; D013025:Glycine max / Q000737:chemistry*; D014357:Trypsin / Q000494:pharmacology; D014358:Trypsin Inhibitor, Bowman-Birk Soybean / Q000494:pharmacology*
chemical_list: D004792:Enzyme Precursors; D061965:Matrix Metalloproteinase Inhibitors; D011480:Protease Inhibitors; D014358:Trypsin Inhibitor, Bowman-Birk Soybean; D014357:Trypsin; D017364:Collagenases; C120487:pro-matrix metalloproteinase 9; D020780:Matrix Metalloproteinase 9; D020781:Matrix Metalloproteinase 1