PDBsum entry 2puq

Hydrolase/hydrolase inhibitor

PDB id: 2puq

Contents

Protein chains

94 a.a. *

254 a.a. *

190 a.a. *

Ligands

TYR-THR-AR7-0QE

BGC

FUC

Metals

_CA

Waters ×187

* Residue conservation analysis

PDB id:

2puq

Name:

Hydrolase/hydrolase inhibitor

Title:

Crystal structure of active site inhibited coagulation factor viia in complex with soluble tissue factor

Structure:

Coagulation factor vii. Chain: l. Fragment: light chain. Synonym: serum prothrombin conversion accelerator. Spca. Proconvertin. Eptacog alfa. Engineered: yes. Coagulation factor vii. Chain: h. Fragment: heavy chain.

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: f7. Expressed in: cricetulus griseus. Expression_system_taxid: 10029. Expression_system_cell: hampster ovary cells. Gene: ace. Expressed in: escherichia coli.

Resolution:

2.05Å R-factor: 0.231 R-free: 0.273

Authors:

J.R.Bjelke,H.B.Rasmussen

Key ref:

K.S.Larsen et al. (2007). Engineering the substrate and inhibitor specificities of human coagulation Factor VIIa. Biochem J, 405, 429-438. PubMed id: 17456045

Date:

09-May-07 Release date: 22-May-07

Supersedes:

2pmm

Protein chain

P08709  (FA7_HUMAN) -  Coagulation factor VII from Homo sapiens

Seq: 466 a.a.

Struc: 94 a.a.

Protein chain

P08709  (FA7_HUMAN) -  Coagulation factor VII from Homo sapiens

Seq: 466 a.a.

Struc: 254 a.a.

Protein chain

P13726  (TF_HUMAN) -  Tissue factor from Homo sapiens

Seq: 295 a.a.

Struc: 190 a.a.

Enzyme reactions

• Enzyme class: Chains L, H: E.C.3.4.21.21 - coagulation factor VIIa.
• Reaction: Hydrolyzes one Arg-|-Ile bond in factor X to form factor Xa.

Biochem J 405:429-438 (2007)

PubMed id: 17456045

Engineering the substrate and inhibitor specificities of human coagulation Factor VIIa.

K.S.Larsen, H.Østergaard, J.R.Bjelke, O.H.Olsen, H.B.Rasmussen, L.Christensen, B.B.Kragelund, H.R.Stennicke.

ABSTRACT

The remarkably high specificity of the coagulation proteases towards macromolecular substrates is provided by numerous interactions involving the catalytic groove and remote exosites. For FVIIa [activated FVII (Factor VII)], the principal initiator of coagulation via the extrinsic pathway, several exosites have been identified, whereas only little is known about the specificity dictated by the active-site architecture. In the present study, we have profiled the primary P4-P1 substrate specificity of FVIIa using positional scanning substrate combinatorial libraries and evaluated the role of the selective active site in defining specificity. Being a trypsin-like serine protease, FVIIa had P1 specificity exclusively towards arginine and lysine residues. In the S2 pocket, threonine, leucine, phenylalanine and valine residues were the most preferred amino acids. Both S3 and S4 appeared to be rather promiscuous, however, with some preference for aromatic amino acids at both positions. Interestingly, a significant degree of interdependence between the S3 and S4 was observed and, as a consequence, the optimal substrate for FVIIa could not be derived directly from a subsite-directed specificity screen. To evaluate the role of the active-site residues in defining specificity, a series of mutants of FVIIa were prepared at position 239 (position 99 in chymotrypsin), which is considered to be one of the most important residues for determining P2 specificity of the trypsin family members. This was confirmed for FVIIa by marked changes in primary substrate specificity and decreased rates of antithrombin III inhibition. Interestingly, these changes do not necessarily coincide with an altered ability to activate Factor X, demonstrating that inhibitor and macromolecular substrate selectivity may be engineered separately.