Expression in Escherichia coli of the human fibrinogen B beta chain and its
cleavage by thrombin
MG Bolyard and ST Lord
Department of Pathology, University of North Carolina, Chapel Hill
27599-7525.
The human fibrinogen B beta chain was expressed in Escherichia coli to
study the functions of fibrinogen associated with this subunit. Recombinant
B beta chains were expressed at 100 ng/mL in an IPTG- dependent manner. A
first cistron sequence, inserted into the expression vector 5' to the B
beta chain cDNA, was required to express the protein. Recombinant B beta
chains were expressed within five minutes after induction with IPTG and
were soluble in physiologic buffers. The recombinant B beta chains migrated
on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) at
a rate identical to B beta chains from fibrinogen treated with N-glycanase.
Recombinant B beta chains were cleaved by thrombin, as demonstrated by the
loss of cross-reactivity with a monoclonal antibody (MoAb) specific for the
undigested B beta 1-42 fragment. The levels of expression of the B beta
chain were much lower than those reported previously for the gamma chain of
fibrinogen expressed in a similar vector in E coli. However, these levels
are sufficient to allow further characterization of this fibrinogen
subunit.
Volume 73,
Issue 5,
pp. 1202-1206,
04/01/1989
Copyright © 1989 by The American Society of Hematology
