title: Isolation and characterization of chicken bile matrix metalloproteinase.
abstract: Avian bile is rich in matrix metalloproteinases (MMP), the enzymes that cleave extracellular matrix proteins such as collagens and proteoglycans. Changes in bile MMP expression have been correlated with hepatic and gall bladder pathologies, but the significance of their expression in normal, healthy bile is not understood. We hypothesized that the MMP in bile may aid the digestion of native collagens that are resistant to conventional gastric proteases. Hence, the objective of this study was to characterize the bile MMP and check its regulation in association with dietary factors. We used substrate zymography, azocoll protease assay, and gelatin affinity chromatography to identify and purify the MMP from chicken bile. Using zymography and SDS PAGE, 5 bands at 70, 64, 58, 50, and 42 kDa were detected. The bands corresponding to 64, 50, and 42 kDa were identified as MMP2 using trypsin in-gel digestion and matrix-assisted laser desorption time-of-flight mass spectrometry and peptide mass fingerprinting. Chickens fed diets containing gelatin supplements showed higher levels of MMP expression in the bile by both azocoll assay and zymography. We conclude that the bile MMP may be associated with the digestion of collagens and other extracellular matrix proteins in avian diets.
authors.affiliation: Cell and Molecular Biology Program, University of Arkansas, Fayetteville 72701 Department of Poultry Science, University of Arkansas, Fayetteville 72701.
authors.forename: B
authors.initials: B
authors.lastname: Packialakshmi
authors.affiliation: Statewide Mass Spectrometry Facility, Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville 72701.
authors.forename: R
authors.initials: R
authors.lastname: Liyanage
authors.affiliation: Department of Poultry Science, University of Arkansas, Fayetteville 72701 USDA, Agricultural Research Service, Poultry Production and Product Safety Research Unit, Fayetteville, AR 72701.
authors.forename: K S
authors.initials: KS
authors.lastname: Rasaputra
authors.affiliation: Statewide Mass Spectrometry Facility, Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville 72701.
authors.forename: Jackson O
authors.initials: JO
authors.lastname: Lay
authors.affiliation: USDA, Agricultural Research Service, Poultry Production and Product Safety Research Unit, Fayetteville, AR 72701 narayan.rath@ars.usda.gov.
authors.forename: N C
authors.initials: NC
authors.lastname: Rath
mesh_terms: D000821:Animal Feed / Q000032:analysis*; D000818:Animals; D001391:Azo Compounds / Q000378:metabolism; D001646:Bile / Q000187:drug effects* / Q000378:metabolism*; D002645:Chickens / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000662:veterinary; D002850:Chromatography, Gel / Q000662:veterinary; D003094:Collagen / Q000378:metabolism; D004032:Diet / Q000662:veterinary; D019587:Dietary Supplements / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel / Q000662:veterinary; D008297:Male; D013058:Mass Spectrometry / Q000662:veterinary; D020782:Matrix Metalloproteinases / Q000302:isolation & purification / Q000378:metabolism*; D011897:Random Allocation
chemical_list: D001391:Azo Compounds; C028470:Azocoll; D003094:Collagen; D020782:Matrix Metalloproteinases
keywords: bile; collagen digestion; mass spectrometry; matrix metalloproteinase-2