 |
PDBsum entry 2a2q
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Hydrolase/blood clotting
|
PDB id
|
|
|
|
2a2q
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
Contents |
 |
|
|
|
|
|
|
|
|
|
|
|
142 a.a.
|
|
|
|
|
|
|
|
|
|
|
254 a.a.
|
|
|
|
|
|
|
|
|
|
|
191 a.a.
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 _CA
×6
|
|
|
|
|
|
|
|
|
|
|
 _CL
×3
|
|
|
|
|
|
|
|
|
|
|
_MG
×3
|
|
|
|
|
|
|
|
|
|
|
 _NA
|
|
|
|
|
|
|
|
|
|
|
_ZN
×2
|
|
|
|
|
|
|
|
|
|
|
|
References listed in PDB file
|
|
|
Key reference
|
|
|
Title
|
|
High resolution structures of p-Aminobenzamidine- And benzamidine-Viia/soluble tissue factor: unpredicted conformation of the 192-193 peptide bond and mapping of ca2+, Mg2+, Na+, And zn2+ sites in factor viia.
|
|
|
Authors
|
|
S.P.Bajaj,
A.E.Schmidt,
S.Agah,
M.S.Bajaj,
K.Padmanabhan.
|
|
|
Ref.
|
|
J Biol Chem, 2006,
281,
24873-24888.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
Abstract
|
|
|
Factor VIIa (FVIIa) consists of a gamma-carboxyglutamic acid (Gla) domain, two
epidermal growth factor-like domains, and a protease domain. FVIIa binds seven
Ca(2+) ions in the Gla, one in the EGF1, and one in the protease domain.
However, blood contains both Ca(2+) and Mg(2+), and the Ca(2+) sites in FVIIa
that could be specifically occupied by Mg(2+) are unknown. Furthermore, FVIIa
contains a Na(+) and two Zn(2+) sites, but ligands for these cations are
undefined. We obtained p-aminobenzamidine-VIIa/soluble tissue factor (sTF)
crystals under conditions containing Ca(2+), Mg(2+), Na(+), and Zn(2+). The
crystal diffracted to 1.8A resolution, and the final structure has an R-factor
of 19.8%. In this structure, the Gla domain has four Ca(2+) and three bound
Mg(2+). The EGF1 domain contains one Ca(2+) site, and the protease domain
contains one Ca(2+), one Na(+), and two Zn(2+) sites. (45)Ca(2+) binding in the
presence/absence of Mg(2+) to FVIIa, Gla-domainless FVIIa, and prothrombin
fragment 1 supports the crystal data. Furthermore, unlike in other serine
proteases, the amide N of Gly(193) in FVIIa points away from the oxyanion hole
in this structure. Importantly, the oxyanion hole is also absent in the
benzamidine-FVIIa/sTF structure at 1.87A resolution. However, soaking
benzamidine-FVIIa/sTF crystals with d-Phe-Pro-Arg-chloromethyl ketone results in
benzamidine displacement, d-Phe-Pro-Arg incorporation, and oxyanion hole
formation by a flip of the 192-193 peptide bond in FVIIa. Thus, it is the
substrate and not the TF binding that induces oxyanion hole formation and
functional active site geometry in FVIIa. Absence of oxyanion hole is unusual
and has biologic implications for FVIIa macromolecular substrate specificity and
catalysis.
|
|
|
|
|
|
|
|
Figure 2.
FIGURE 2. Conformation of the  loop and positions of
the Ca^2^+ and Mg^2^+ ions in the FVIIa Gla domain. A,
superpositioning of the Gla domain in the presence of
Ca^2+/Mg^2+ versus Ca^2+ (19). The C  atoms used for
superpositioning were residues 13L-46L. The Ca^2+/Mg^2+
structure is in blue and the Ca^2+ structure is in magenta.
Phe^4, Leu^5, Gla6, Gla7, and Leu^8 are depicted for both
structures. Ca^2+ (blue) and Mg^2+ (cyan) ions for the present
structure as well as Ca^2+ ions (magenta) for the PDB code 1DAN
(19) are shown as spheres. N represents the N terminus of the
Gla domain of FVIIa. B, coordination of the Ca^2+ and Mg^2+ ions
in the Gla domain of the Ca^2+/Mg^2+ structure. Electron density
(2F[obs] - F[calc]) contoured at 1  of all nine Gla
residues as well as that of Ca^2+ (magenta spheres), Mg^2+ (cyan
spheres), and coordinating water molecules (red spheres)is
shown. Coordination of Ca5 to O of Ala^1L and H-bonds between
Gla6 and the NH[2] side chain of Arg^9L, Gla7, and NofPhe^4L,
and a water molecule with O  [1] of Gln^2L are
indicated with dashed arrows. Black dashed lines depict all
other coordinations and H-bonds. A stereo figure is given in the
supplemental material as Fig. 1S.
|
|
Figure 5.
FIGURE 5. Ca^2^+ site in the EGF1 domain and location of
the Zn^2^+ sites and their linkage to the protease domain Ca^2^+
site in FVIIa. A, Ca^2+ site in the EGF1 domain. Electron
density (2F[obs] - F[calc]) contoured at 1 (gray) for Ca^2+
(magenta sphere) and two water molecules (red spheres) is shown.
Electron density contoured at 5 for Ca^2+ is shown in
blue. Note that in the presence of Ca^2+, Mg^2+ will not occupy
this site. All eight coordination ligands (black dashed lines)
are shown, and the residues labeled are those of the light chain
of FVIIa. B, location of the Zn^2+ sites and their linkage to
the protease domain Ca^2+ site. Electron density (2F[obs] -
F[calc]) contoured at 1 (gray) of Zn^2+ (cyan
spheres), Ca^2+ (magenta sphere), and water molecules (red
spheres) is shown. The electron density contoured at 3 for
Zn^2+ and Ca^2+ ions is shown in blue. Note the linkage between
the Zn^2+ sites and the Ca^2+ site. The metal ion coordination
to its ligands and H-bonds between water molecules is shown with
black dotted lines. Residue numbering used in the figure is that
of chymotrypsin.
|
|
|
|
|
|
The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2006,
281,
24873-24888)
copyright 2006.
|
|
|
|
|
|
|
