Posttranslational modifications and activity of natural and recombinant tissue factor.
Tissue factor is a membrane protein, which in a complex with factor VIIa initiates in vivo blood coagulation. Due to the scarcity of natural tissue factor protein, most studies have relied upon recombinant tissue factor forms. However, there have been only cursory experimental comparisons of natural and recombinant tissue factor proteins. Our preliminary data suggested that placental tissue factor in a complex with factor VIIa was more efficient activator of factor X than the recombinant protein. After deglycosylation, both forms of tissue factor showed almost an identical activity in the extrinsic factor Xase. Analyses using tryptic digestion and mass-spectrometry revealed that the levels of glycosylation and the composition of carbohydrates present in natural placental tissue factor were different than those in its recombinant counterpart. These data indicate that natural and recombinant tissue factor proteins differ in their posttranslational modifications and that these differences translate into different cofactor activity. Thus the use of recombinant tissue factor proteins for the quantitation of natural tissue factor is misleading.
Butenas
Saulius
S
Department of Biochemistry, University of Vermont, 208 South Park Drive, Burlington, VT 05446, USA. sbutenas@uvm.edu
Krudysz-Amblo
Jolanta
J
Mann
Kenneth G
KG
D001777:Blood Coagulation; D002241:Carbohydrates / Q000737:chemistry; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D015942:Factor VIIa / Q000737:chemistry; D006023:Glycoproteins / Q000737:chemistry; D006031:Glycosylation; D006801:Humans; D008358:Mannose / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods; D008970:Molecular Weight; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000737:chemistry*; D013925:Thromboplastin / Q000737:chemistry*
D002241:Carbohydrates; D006023:Glycoproteins; D011994:Recombinant Proteins; D013925:Thromboplastin; D015942:Factor VIIa; D008358:Mannose