PDBsum entry 1no9

Hydrolase/hydrolase inhibitor

PDB id: 1no9

Contents

Protein chains

36 a.a. *

253 a.a. *

11 a.a. *

Ligands

NAG

4ND

Waters ×147

* Residue conservation analysis

PDB id:

1no9

Name:

Hydrolase/hydrolase inhibitor

Title:

Design of weakly basic thrombin inhibitors incorporating novel p1 binding functions: molecular and x-ray crystallographic studies.

Structure:

Alpha thrombin. Chain: l. Fragment: light chain. Alpha thrombin. Chain: h. Fragment: heavy chain. Hirugen. Chain: i. Fragment: chain i.

Source:

Homo sapiens. Human. Organism_taxid: 9606. Other_details: plasma. Synthetic: yes. Hirudo medicinalis. Organism_taxid: 6421

Biol. unit:

Trimer (from PQS)

Resolution:

1.90Å R-factor: 0.183 R-free: 0.204

Authors:

G.De Simone,V.Menchise,S.Omaggio,C.Pedone,A.Scozzafava,C.T.Supuran

Key ref:

G.De Simone et al. (2003). Design of weakly basic thrombin inhibitors incorporating novel P1 binding functions: molecular and X-ray crystallographic studies. Biochemistry, 42, 9013-9021. PubMed id: 12885234 DOI: 10.1021/bi020512l

Date:

16-Jan-03 Release date: 26-Aug-03

Protein chain

P00734  (THRB_HUMAN) -  Prothrombin from Homo sapiens

Seq: 622 a.a.

Struc: 36 a.a.

Protein chain

P00734  (THRB_HUMAN) -  Prothrombin from Homo sapiens

Seq: 622 a.a.

Struc: 253 a.a.

Protein chain

P01050  (HIRV1_HIRME) -  Hirudin variant-1 from Hirudo medicinalis

Seq: 65 a.a.

Struc: 11 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: Chains L, H: E.C.3.4.21.5 - thrombin.
• Reaction: Preferential cleavage: Arg-|-Gly; activates fibrinogen to fibrin and releases fibrinopeptide A and B.

DOI no: 10.1021/bi020512l

Biochemistry 42:9013-9021 (2003)

PubMed id: 12885234

Design of weakly basic thrombin inhibitors incorporating novel P1 binding functions: molecular and X-ray crystallographic studies.

G.De Simone, V.Menchise, S.Omaggio, C.Pedone, A.Scozzafava, C.T.Supuran.

ABSTRACT

To prepare weakly basic thrombin inhibitors with modified S1 anchoring groups, two series of compounds were synthesized by reaction of guanidine or aminoguanidine with acyl halides and N,N-disubstituted carbamoyl chlorides. pK(a) measurements of these acylated guanidines/aminoguanidines showed a reduced basicity, with pK(a) values in the range of 8.4-8.7. These molecules typically showed inhibition constants in the range of 150-425 nM against thrombin and 360-965 nM against trypsin, even though some bulky derivatives, such as N,N-diphenylcarbamoylguanidine/aminoguanidine and their congeners, showed much stronger thrombin inhibitory activity, with inhibition constants in the range of 24-42 nM. Unexpectedly, very long incubation times with both proteases revealed that aminoguanidine derivatives behaved as irreversible inhibitors. To assess the molecular basis responsible for the high affinity observed for these molecules toward thrombin, the crystal structure of the thrombin-hirugen-N,N-diphenylcarbamoylaminoguanidine complex has been solved at 1.90 A resolution. The structural analysis of the complex revealed an unexpected interaction mode with the protease, resulting in an N,N-diphenylcarbamoyl intermediate covalently bound to the catalytic serine as a consequence of its hydrolysis together with the release of the aminoguanidine moiety. Surprisingly, in this covalent adduct a phenyl group was found in the S1 specificity pocket, which usually recognizes positively charged residues. These findings provide new insights in the design of low basicity serine protease inhibitors.