title: Clotting of mammalian fibrinogens by papain: a re-examination.
abstract: Papain has long been known to cause the gelation of mammalian fibrinogens. It has also been reported that papain-fibrin is insoluble in dispersing solvents like strong urea or sodium bromide solutions, similar to what is observed with thrombin-generated clots in the presence of factor XIIIa and calcium. In those old studies, both the gelation and subsequent clot stabilization were attributed to papain, although the possibility that the second step might be due to contaminating factor XIII in fibrinogen preparations was considered. I have revisited this problem in light of knowledge acquired over the past half-century about thiol proteases like papain, which mostly cleave peptide bonds, and transglutaminases like factor XIIIa that catalyze the formation of ε-lysyl-γ-glutamyl cross-links. Recombinant fibrinogen, inherently free of factor XIII and other plasma proteins, formed a stable gel when treated with papain alone. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that the intermolecular cross-linking in papain-fibrin leads to γ-chain dimers, trimers, and tetramers, just as is the case with thrombin-factor XIIIa-stabilized fibrin. Mass spectrometry of bands excised from gels showed that the cross-linked material is quite different from what occurs with factor XIIIa, however. With papain, the cross-linking occurs between γ chains in neighboring protofibrils becoming covalently linked in a "head-to-tail" fashion by a transpeptidation reaction involving the α-amino group of γ-Tyr1 and a papain cleavage site at γ-Gly403 near the carboxy terminus, rather than by the (reciprocal) "tail-to-tail" manner that occurs with factor XIIIa and that depends on cross-links between γ-Lys406 and γ-Gln398.
authors.affiliation: Departments of Chemistry & Biochemistry and Molecular Biology, University of California at San Diego , La Jolla, California 92093-0314, United States.
authors.forename: Russell F
authors.initials: RF
authors.lastname: Doolittle
mesh_terms: D002403:Cathepsins / Q000737:chemistry*; D003432:Cross-Linking Reagents / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D005176:Factor XIII / Q000737:chemistry / Q000235:genetics; D005340:Fibrinogen / Q000737:chemistry*; D006801:Humans; D010206:Papain / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D013997:Time Factors
chemical_list: D003432:Cross-Linking Reagents; D011994:Recombinant Proteins; D005340:Fibrinogen; D005176:Factor XIII; D002403:Cathepsins; D010206:Papain; C028217:cathepsin S