850
c h a p t e r 36
Biochemistry of Hemostasis
F I G U R E 3 6 - 7
(Also see color figure.) A group of structurally similar protein inhibitors of
the serine proteinases known as SERPINS (SERine Proteinase INhibitors).
The structure shown is human antithrombin. The reference SERPIN,
№
i -proteinase inhibitor or
a
i -antitrypsin, contains ~30% a helix
(9 helices) and 40% sheet (5
ft
sheets). Other members of the SERPIN
family contain both additional helices and
ft
sheets. The reactive center
loop of antithrombin, residues 378-396, contains the reactive site residues
Arg393 and Ser394. Upon reaction with the target proteinase or after
cleavage by the target proteinase (a reaction that inactivates the inhibitor
without inactivating the proteinase), the reactive center loop folds between
the S3 and S5 sheets.
(SERine Proteinase INhibitors) (Figure 36-7). There
are three SERPINS that inactivate proteinases in the
procoagulant subsystem, one in the anticoagulant subsys-
tem, and three in the fibrinolytic subsystem. Regulation
of the action of the proteinases requires a mechanism
compatible with the fact that proteinase precursor ac-
tivation is irreversible. This is achieved by the protein
proteinase inhibitors acting as suicide substrates, i.e.,
the reaction between proteinase and inhibitor proceeds
“half-way” when compared with the proteolytic reaction.
The reaction between proteinase and inhibitor stops at the
stage of acyl enzyme or tetrahedral intermediate. Water
normally does reach the active site in the proteinase
substrate complex to achieve hydrolysis, but the serpin
prevents this, and thus the proteinase is “killed” by the
inhibitor. All SERPINS react with their target proteinases
irreversibly and form
1 : 1
stoichiometric complexes.
The reaction of the proteinase with the inhibitor occurs
at a basic residue (Arg or Lys residue) in a loop that ex-
tends away from the globular inhibitor molecule. This ba-
sic residue, called the reactive site residue, is locked in the
proteinase active site as the acyl enzyme. The reactive site
and the complete amino acid sequences are similar in all
SERPINS.
Other Proteins of the Hemostatic System
Proteins that do not fit into the previous functional cate-
gories are fibrinogen, factor XIII (the plasma protransglu-
taminase), and von Willebrand factor.
Fibrinogen
Fibrinogen is a precursor to the spon-
taneously polymerizing protein fibrin. Fibrin is the sub-
stance of the gelatinous clot and, thus, fibrinogen and fibrin
are best categorized as structural proteins. Fibrin strands,
as noted above, provide the reinforcement necessary for
an adequate hemostatic plug. Fibrin can also act as a “sur-
face” for fibrinolytic system proteins. This latter function
serves to localize fibrinolysis to the fibrin clot. The struc-
ture of fibrinogen, as deduced from electron microscopy
and other techniques, is shown in schematic form in
Figure 36-8.
Factor XIII (Plasma Protransglutaminase)
Factor
XIII, after activation to factor Xllla, is a transglutami-
nase that catalyzes the formation of covalent cross-links
in fibrin. In the absence of its action, the fibrin structure is
unstable and physiologically inadequate. Factor XIII ex-
ists in two forms, as a tetramer in plasma, a
2
b
2
, and as a
dimer, a
2
, in the platelet granules. Activated factor XIII cat-
alyzes the formation of “isopeptide,” £-(y-glutamyl (lysine
bonds between the e-amino groups of Lys residues and
y-carboxamido groups of Gin residues of fibrin monomers
in polymerized fibrin. The a subunits contain the active site
residues that are directly involved in the transamidation
reaction. The a
2
b
2
subunits dissociate in the presence of
Ca2+ to form a
2
and b
2
dimers. In plasma, which contains
~ 1 .5 mM Ca2+, free a
2
and free b
2
subunits are found in
addition to the tetramer.
Von
Willebrand
Factor
Von
Willebrand
factor
(vWF) is a multisubunit protein that serves both to an-
chor the platelets to the subendothelium and as a car-
rier protein for factor VIII in the plasma. The circulating
vWF is the largest protein of the hemostatic system. It is
made of protomeric units which themselves are dimers of
the 250,000-Da vWF polypeptide chain. The vWF gene
product is a single polypeptide chain that associates to
form the dimeric protomer. Disulfide bonds link the two
vWF monomer chains in the protomer. The 500,000-Da
dimeric protomers further associate to form the more than
20,000,000-Da vWF molecules present in the circulation.
Recognition sites are present on the von Willebrand factor
