title: Prolyl 4-hydroxylase in the foot of the marine mussel Mytilus edulis L.: purification and characterization.
abstract: The mussel foot secretes a variety of unusual hydroxyproline-containing collagenous and noncollagenous proteins. Prolyl 4-hydroxylase acting on one or more of the secreted proteins was isolated from the foot by using conventional gel filtration and ion exchange chromatography. Mr of the intact enzyme was 230,000 (alpha 2 beta 2) composed of two subunits with Mr of 60,000 (alpha) and 57,000 (beta) as estimated by HPLC gel filtration and SDS-PAGE. The enzyme utilized (Pro-Pro-Gly)10 as a substrate with an apparent Km value of 0.17 mM. Cofactors and inhibitors were very similar to animal, plant, and microbial prolyl hydroxylases previously described. The enzyme had a relatively sharp pH optimum in the range of 7.8-8.3 and the hydroxyproline formed increased in proportion to the rise in the temperature between 5 and 20 degrees C. No detectable hydroxylation occurred with poly-L-proline or the unhydroxylated decapeptide analog (Ala-Lys-Pro-Ser-Tyr-Pro-Pro-Thr-Tyr-Lys) of the polyphenolic protein. Kinetic studies, however, revealed that the mussel prolyl 4-hydroxylase was competitively inhibited by poly-L-proline and uncompetitively inhibited by the decapeptide. These results suggest that the decapeptide binds the enzyme-substrate i.e. (Pro-Pro-Gly)10 complex. It is not yet clear whether this enzyme acts exclusively on collagenous substrates or whether its catalytic purview extends as well to the polyphenolic protein.
authors.affiliation: College of Marine Studies, University of Delaware, Lewes 19958.
authors.forename: K
authors.initials: K
authors.lastname: Marumo
authors.forename: J H
authors.initials: JH
authors.lastname: Waite
mesh_terms: D000818:Animals; D049872:Bivalvia / Q000201:enzymology*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D011393:Procollagen-Proline Dioxygenase / Q000032:analysis / Q000302:isolation & purification*
chemical_list: D011393:Procollagen-Proline Dioxygenase