title: Profactor IX: the propeptide inhibits binding to membrane surfaces and activation by factor XIa.
abstract: The gamma-carboxylase recognition site in the propeptide of profactor IX signals the gamma-carboxylation of specific glutamic acid residues in the adjacent Gla domain during factor IX biosynthesis. To study posttranslational processing of the vitamin K-dependent blood coagulation factors and the properties of processing intermediates, we have isolated an incompletely processed factor IX species, profactor IX, from the medium of heterologous mammalian cells expressing the human factor IX cDNA. Profactor IX was purified by sequential immunoaffinity chromatography using antibodies specific for the propeptide and antibodies specific for the well-carboxylated factor IX species. This purified profactor IX preparation was fully gamma-carboxylated and contained the N-terminal propeptide, but it exhibited no factor IX procoagulant activity. Profactor IX was not cleaved following incubation with factor XIa. In contrast to mature factor IX, profactor IX did not demonstrate Ca(II)-dependent binding to acidic phospholipid vesicles, nor can the membrane binding surface be expressed, as detected by antibodies specific for this epitope. The propeptide of profactor IX can be removed in vitro by a specific endopeptidase, furin/PACE, yielding factor IX, which can be converted to fully active factor IXa by factor XIa and which binds normally to acidic phospholipid vesicles. These results indicate that fully gamma-carboxylated profactor IX is biologically inactive due to the presence of the propeptide.
authors.affiliation: Department of Biochemistry, Tufts University School of Medicine, Boston, Massachusetts.
authors.forename: J A
authors.initials: JA
authors.lastname: Bristol
authors.forename: S J
authors.initials: SJ
authors.lastname: Freedman
authors.forename: B C
authors.initials: BC
authors.lastname: Furie
authors.forename: B
authors.initials: B
authors.lastname: Furie
mesh_terms: D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D016466:CHO Cells; D002118:Calcium / Q000494:pharmacology; D002262:Carboxy-Lyases / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D006224:Cricetinae; D004492:Edetic Acid / Q000494:pharmacology; D005164:Factor IX / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology*; D015945:Factor XIa / Q000494:pharmacology*; D006801:Humans; D008081:Liposomes / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism; D011487:Protein Conformation; D011498:Protein Precursors / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000302:isolation & purification
chemical_list: D008081:Liposomes; D010743:Phospholipids; D011498:Protein Precursors; D011994:Recombinant Proteins; D005164:Factor IX; D004492:Edetic Acid; D015945:Factor XIa; D002262:Carboxy-Lyases; D002118:Calcium