|
|
|
|
|
 |
Contents |
 |
|
|
|
|
|
|
|
|
|
|
|
|
* Residue conservation analysis
|
|
|
|
|
|
|
|
|
|
Enzyme class:
|
|
Chains A, B, C, D:
E.C.3.4.21.5
- thrombin.
|
|
|
|
|
|
|
Reaction:
|
|
Preferential cleavage: Arg-|-Gly; activates fibrinogen to fibrin and releases fibrinopeptide A and B.
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
|
|
|
|
|
|
| |
|
DOI no:
|
Biochemistry
37:6645-6657
(1998)
|
|
PubMed id:
|
|
|
|
|
|
| |
|
Novel natural product 5,5-trans-lactone inhibitors of human alpha-thrombin: mechanism of action and structural studies.
|
|
M.P.Weir,
S.S.Bethell,
A.Cleasby,
C.J.Campbell,
R.J.Dennis,
C.J.Dix,
H.Finch,
H.Jhoti,
C.J.Mooney,
S.Patel,
C.M.Tang,
M.Ward,
A.J.Wonacott,
C.W.Wharton.
|
|
|
|
|
| |
ABSTRACT
|
|
|
|
| |
|
|
High-throughput screening of methanolic extracts from the leaves of the plant
Lantana camara identified potent inhibitors of human alpha-thrombin, which were
shown to be 5,5-trans-fused cyclic lactone euphane triterpenes [O'Neill et al.
(1998) J. Nat. Prod. (submitted for publication)]. Proflavin displacement
studies showed the inhibitors to bind at the active site of alpha-thrombin and
alpha-chymotrypsin. Kinetic analysis of alpha-thrombin showed tight-binding
reversible competitive inhibition by both compounds, named GR133487 and
GR133686, with respective kon values at pH 8.4 of 1.7 x 10(6) s-1 M-1 and 4.6 x
10(6) s-1 M-1. Electrospray ionization mass spectrometry of thrombin/inhibitor
complexes showed the tight-bound species to be covalently attached, suggesting
acyl-enzyme formation by reaction of the active-site Ser195 with the
trans-lactone carbonyl. X-ray crystal structures of alpha-thrombin/GR133686 (3.0
A resolution) and alpha-thrombin/GR133487 (2.2 A resolution) complexes showed
continuous electron density between Ser195 and the ring-opened lactone carbonyl,
demonstrating acyl-enzyme formation. Turnover of inhibitor by alpha-thrombin was
negligible and mass spectrometry of isolated complexes showed that reversal of
inhibition occurs by reformation of the trans-lactone from the acyl-enzyme. The
catalytic triad appears undisrupted and the inhibitor carbonyl occupies the
oxyanion hole, suggesting the observed lack of turnover is due to exclusion of
water for deacylation. The acyl-enzyme inhibitor hydroxyl is properly positioned
for nucleophilic attack on the ester carbonyl and therefore relactonization;
furthermore, the higher resolution structure of alpha-thrombin/GR133487 shows
this hydroxyl to be effectively superimposable with the recently proposed
deacylating water for peptide substrate hydrolysis [Wilmouth, R. C., et al.
(1997) Nat. Struct.Biol. 4, 456-462], suggesting the alpha-thrombin/GR133487
complex may be a good model for this reaction.
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
 |
 |
|
Literature references that cite this PDB file's key reference
|
|
|
| |
PubMed id
|
|
Reference
|
|
|
|
|
|
A.D.Borthwick
(2005).
Design of translactam HCMV protease inhibitors as potent antivirals.
|
| |
Med Res Rev,
25,
427-452.
|
|
|
|
|
|
|
S.Srivastava,
L.N.Goswami,
and
D.K.Dikshit
(2005).
Progress in the design of low molecular weight thrombin inhibitors.
|
| |
Med Res Rev,
25,
66-92.
|
|
|
|
|
|
|
G.Müller
(2003).
Medicinal chemistry of target family-directed masterkeys.
|
| |
Drug Discov Today,
8,
681-691.
|
|
|
|
|
|
|
B.Doljak,
M.Stegnar,
U.Urleb,
S.Kreft,
A.Umek,
M.Ciglaric,
B.Strukelj,
and
T.Popovic
(2001).
Screening for selective thrombin inhibitors in mushrooms.
|
| |
Blood Coagul Fibrinolysis,
12,
123-128.
|
|
|
|
|
|
|
H.Jhoti,
A.Cleasby,
S.Reid,
P.J.Thomas,
M.Weir,
and
A.Wonacott
(1999).
Crystal structures of thrombin complexed to a novel series of synthetic inhibitors containing a 5,5-trans-lactone template.
|
| |
Biochemistry,
38,
7969-7977.
|
|
|
PDB codes:
|
|
|
|
|
|
|
|
M.Pass,
R.E.Bolton,
S.J.Coote,
H.Finch,
S.Hindley,
A.Lowdon,
E.McDonald,
J.McLaren,
M.Owen,
N.A.Pegg,
C.J.Mooney,
C.M.Tang,
S.Parry,
and
C.Patel
(1999).
Synthetic [5,5] trans-fused indane lactones as inhibitors of thrombin.
|
| |
Bioorg Med Chem Lett,
9,
431-436.
|
|
|
|
|
|
|
R.C.Wilmouth,
S.Kassamally,
N.J.Westwood,
R.J.Sheppard,
T.D.Claridge,
R.T.Aplin,
P.A.Wright,
G.J.Pritchard,
and
C.J.Schofield
(1999).
Mechanistic insights into the inhibition of serine proteases by monocyclic lactams.
|
| |
Biochemistry,
38,
7989-7998.
|
|
|
PDB code:
|
|
|
|
|
|
|
|
R.C.Wilmouth,
N.J.Westwood,
K.Anderson,
W.Brownlee,
T.D.Claridge,
I.J.Clifton,
G.J.Pritchard,
R.T.Aplin,
and
C.J.Schofield
(1998).
Inhibition of elastase by N-sulfonylaryl beta-lactams: anatomy of a stable acyl-enzyme complex.
|
| |
Biochemistry,
37,
17506-17513.
|
|
|
PDB code:
|
|
|
|
|
|
|
The most recent references are shown first.
Citation data come partly from CiteXplore and partly
from an automated harvesting procedure. Note that this is likely to be
only a partial list as not all journals are covered by
either method. However, we are continually building up the citation data
so more and more references will be included with time.
Where a reference describes a PDB structure, the PDB
codes are
shown on the right.
|
 |
Links
