Factor VIIa and the extracellular domains of human tissue factor form a compact complex: A study by X‐ray and neutron solution scattering

Ashton, Alun; Kemball‐Cook, Geoffrey; Johnson, Dwayne; Martin, David; O'Brien, Donogh P.; Tuddenham, Edward G. D.; Perkins, Stephen J.

doi:10.1016/0014-5793(95)01093-t

Cited by 21 publications

(31 citation statements)

References 27 publications

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“…The relative orientation of domains is speculative except when multiple close contacts are defined, such as between the second EGF-like and the protease domains. An extended structure of fVIIa has been inferred from X-ray and neutron scattering data (Ashton et al, 1995), but unpublished observations in our laboratory indicate a possible interaction between the Gla and protease domains.…”

Section: Discussionmentioning

confidence: 89%

Structural changes in factor VIIa induced by Ca²⁺ and tissue factor studied using circular dichroism spectroscopy

1996

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Factor VIIa (fVIIa) is composed of four discrete domains, a y-carboxyglutamic acid (G1a)-containing domain, two epidermal growth factor (EGF)-like domains, and a serine protease domain, all of which appear to be involved, to different extents, in an optimal interaction with tissue factor (TF). All except the second EGF-like domain contain at least one Ca2+ binding site and many properties of fVIIa, e.g., TF and phospholipid binding and amidolytic activity, are Ca2+-dependent. A CD study was performed to characterize and locate the conformational changes in fVIIa induced by Ca'+ and TF binding. In addition to intact fVIIa, derivatives lacking the Gla domain or the protease domain were used. Assignment of the Ca2+-induced changes in the far-UV region of the fVIIa spectrum to the Gla domain could be made by comparing the CD spectra obtained with these fVIla derivatives. The changes primarily appeared to reflect a Ca*+-induced ordering of a-helices existing in the apo state of fVIIa. This was corroborated by models of the apo and Ca2+ forms of fVIIa constructed on the basis of known structures of homologous proteins. Far-UV spectra of the Gla domain of fVIIa, obtained as difference spectra between fVIIa derivatives, were very similar to those of isolated Gla peptides from other vitamin K-dependent plasma proteins. The near-UV CD spectrum of fVIIa was dominated by aromatic residues residing in the protease domain and specific bands affected by Ca2+ were indicative of tertiary structural alterations. The formation of a NIIa:TF complex led to secondary structural changes that appeared to be restricted to the catalytic domain, possibly shedding light on the mechanism by which T F induces an enhancement of fVIIa catalytic activity.

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Section: Discussionmentioning

confidence: 89%

Structural changes in factor VIIa induced by Ca²⁺ and tissue factor studied using circular dichroism spectroscopy

1996

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“…It has been reported that the WTFVIIa molecule free in solution has an extended conformation (38) and possesses interdomain flexibility (resulting from Gla-EGF1 and EGF1-EGF2 flexibility) which is reduced on binding to sTF (39). It is likely that both the high affinity of the sTF-WTFVIIa interaction and part of the allosteric promotion of FVIIa activity which results derive from the stabilization by TF of FVIIa inter-and intradomain conformation, requiring some movement of FVIIa domains to orient the large surface involved.…”

Section: Discussionmentioning

confidence: 99%

Coagulation Factor VII Gln100 → Arg

Kemball‐Cook¹,

Johnson²,

Takahashi³

et al. 1998

Journal of Biological Chemistry

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We have used recombinant mammalian expression and purification of the factor VII (FVII) variant Gln 100 3 Arg (Q100RFVII) to study FVII deficiency in subjects with this mutation. Q100RFVII was secreted poorly in comparison with wild-type FVII (WTFVII) in a stable mammalian expression system, and purified variant protein was found to have undetectable clotting activity. Following activation by immobilized factor Xa, Q100RFVIIa had amidolytic activity similar to WTFVIIa in the absence of soluble tissue factor (sTF); however, unlike WTFVIIa no typical increase in activity was seen after addition of sTF. In a factor X activation assay using relipidated transmembrane truncated tissue factor (residues 1-243), Q100RFVIIa showed less than 5% of the ability of WTFVIIa to activate factor X.We performed direct binding analysis of WT and Q100RFVII/FVIIa to immobilized sTF using surface plasmon resonance, and severely reduced binding of both non-activated and activated Q100RFVII to sTF was seen, indicating a pronounced defect in tissue factor (TF) interaction with this variant.In the sTF-FVIIa crystal structure the candidate residue Gln 100 is not in contact with TF but is at the epidermal growth factor 2-protease domain interface. We suggest that the mutation results in a global fold change severely reducing tissue factor interaction; mutation of FVII residues not directly involved in the interaction with TF may still result in variant FVII unable to take part in the initiation of coagulation.

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“…However, the different techniques in these studies yielded different conclusions about the solution structure of the sTF⅐fVIIa complex. In one study, the proteins appeared to be positioned side-by-side in the complex (6), while in the other study, the complex appeared to be substantially more elongated than either of its components (5). Crystal structures of both sTF (7,8) and sTF⅐fVIIa (9) have recently been determined, and in the crystals, sTF and fVIIa are elongated and aligned side-by-side in the sTF⅐fVIIa complex (7)(8)(9).…”

mentioning

confidence: 97%

“…The solution structures of both fVIIa and sTF, the soluble extracellular domain of TF, have been shown, using various biophysical techniques, to be asymmetric or elongated (5,6). However, the different techniques in these studies yielded different conclusions about the solution structure of the sTF⅐fVIIa complex.…”

mentioning

confidence: 99%

The Location of the Active Site of Blood Coagulation Factor VIIa above the Membrane Surface and Its Reorientation upon Association with Tissue Factor

McCallum

Hapak

Neuenschwander

et al. 1996

Journal of Biological Chemistry

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The topography of membrane-bound blood coagulation factor VIIa (fVIIa) was examined by positioning a fluorescein dye in the active site of fVIIa via a tripeptide tether to yield fluorescein-D-phenylalanyl-L-prolyl-Larginyl-fVIIa (Fl-FPR-fVIIa). The location of the activesite probe relative to the membrane surface was determined, both in the presence and absence of tissue factor (TF), using fluorescence energy transfer between the fluorescein dye and octadecylrhodamine (OR) at the phospholipid vesicle surface. When Fl-FPR-fVIIa was titrated with phospholipid vesicles containing OR, the magnitude of OR-, calcium ion-, and phosphatidylserinedependent fluorescence energy transfer revealed that the average distance of closest approach between fluorescein in the active site of fVIIa and OR at the vesicle surface is 82 Å assuming a random orientation of donor and acceptor dyes ( 2 ‫؍‬ 2/3; the orientational uncertainty totals ϳ10%). The active site of fVIIa is therefore located far above the membrane surface, and the elongated fVIIa molecule must bind at one end to the membrane and project approximately perpendicularly out of the membrane.When Fl-FPR-fVIIa was titrated with vesicles that contained TF, the efficiency of energy transfer was increased by a TF-dependent translational and/or rotational movement of the fVIIa protease domain relative to the membrane surface. If this movement was solely translational, the height of the active site of fVIIa was lowered by an average of 6 Å after binding to TF. The association of fVIIa with TF on the membrane surface therefore causes a significant reorientation of the active site relative to the membrane surface. This cofactor-dependent realignment of the active-site groove presumably facilitates and optimizes fVIIa cleavage of its membrane-bound substrates.

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Factor VIIa and the extracellular domains of human tissue factor form a compact complex: A study by X‐ray and neutron solution scattering

Cited by 21 publications

References 27 publications

Structural changes in factor VIIa induced by Ca²⁺ and tissue factor studied using circular dichroism spectroscopy

Structural changes in factor VIIa induced by Ca²⁺ and tissue factor studied using circular dichroism spectroscopy

Coagulation Factor VII Gln100 → Arg

The Location of the Active Site of Blood Coagulation Factor VIIa above the Membrane Surface and Its Reorientation upon Association with Tissue Factor

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Factor VIIa and the extracellular domains of human tissue factor form a compact complex: A study by X‐ray and neutron solution scattering

Cited by 21 publications

References 27 publications

Structural changes in factor VIIa induced by Ca2+ and tissue factor studied using circular dichroism spectroscopy

Structural changes in factor VIIa induced by Ca2+ and tissue factor studied using circular dichroism spectroscopy

Coagulation Factor VII Gln100 → Arg

The Location of the Active Site of Blood Coagulation Factor VIIa above the Membrane Surface and Its Reorientation upon Association with Tissue Factor

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Structural changes in factor VIIa induced by Ca²⁺ and tissue factor studied using circular dichroism spectroscopy

Structural changes in factor VIIa induced by Ca²⁺ and tissue factor studied using circular dichroism spectroscopy