Crystal structures of the FXIa catalytic domain in complex with ecotin mutants reveal substrate-like interactions

被引:46
作者
Jin, L [1 ]
Pandey, P [1 ]
Babine, RE [1 ]
Gorga, JC [1 ]
Seidl, KJ [1 ]
Gelfand, E [1 ]
Weaver, DT [1 ]
Abdel-Meguid, SS [1 ]
Strickler, JE [1 ]
机构
[1] Daiichi Asubio Med Res Labs LLC, Cambridge, MA 02139 USA
关键词
D O I
10.1074/jbc.M411309200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Thrombosis can lead to life-threatening conditions such as acute myocardial infarction, pulmonary embolism, and stroke. Although commonly used anti-coagulant drugs, such as low molecular weight heparin and warfarin, are effective, they carry a significant risk of inducing severe bleeding complications, and there is a need for safer drugs. Activated Factor XI (FXIa) is a key enzyme in the amplification phase of the coagulation cascade. Anti-human FXI antibody significantly reduces thrombus growth in a baboon thrombosis model without bleeding problems (Gruber, A., and Hanson, S. R. (2003) Blood 102, 953-955). Therefore, FXIa is a potential target for anti-thrombosis therapy. To determine the structure of FXIa, we derived a recombinant catalytic domain of FXI, consisting of residues 370-607 (rhFXI(370-607)). Here we report the first crystal structure of rhFXI(370-607) in complex with a substitution mutant of ecotin, a panserine protease protein inhibitor secreted by Escherichia coli, to 2.2 Angstrom resolution. The presence of ecotin not only assisted in the crystallization of the enzyme but also revealed unique structural features in the active site of FXIa. Subsequently, the sequence from P5 to P2' in ecotin was mutated to the FXIa substrate sequence, and the structures of the rhFXI(370-607)-ecotin mutant complexes were determined. These structures provide us with an understanding of substrate binding interactions of FXIa, the structural information essential for the structure-based design of FXIa-selective inhibitors.
引用
收藏
页码:4704 / 4712
页数:9
相关论文
共 48 条
[21]   Engineering inhibitors highly selective for the S1 sites of Ser190 trypsin-like serine protease drug targets [J].
Katz, BA ;
Sprengeler, PA ;
Luong, C ;
Verner, E ;
Elrod, K ;
Kirtley, M ;
Janc, J ;
Spencer, JR ;
Breitenbucher, JG ;
Hui, H ;
McGee, D ;
Allen, D ;
Martelli, A ;
Mackman, RL .
CHEMISTRY & BIOLOGY, 2001, 8 (11) :1107-1121
[22]   Structural basis for selectivity of a small molecule, S1-binding, submicromolar inhibitor of urokinase-type plasminogen activator [J].
Katz, BA ;
Mackman, R ;
Luong, C ;
Radika, K ;
Martelli, A ;
Sprengeler, PA ;
Wang, J ;
Chan, HD ;
Wong, L .
CHEMISTRY & BIOLOGY, 2000, 7 (04) :299-312
[23]   What can the structures of enzyme-inhibitor complexes tell us about the structures of enzyme substrate complexes? [J].
Laskowski, M ;
Qasim, MA .
BIOCHIMICA ET BIOPHYSICA ACTA-PROTEIN STRUCTURE AND MOLECULAR ENZYMOLOGY, 2000, 1477 (1-2) :324-337
[24]   HEREDITY AND COAGULATION STUDIES IN 10 FAMILIES WITH FACTOR 11 (PLASMA THROMBOPLASTIN ANTECEDENT) DEFICIENCY [J].
LEIBA, H ;
RAMOT, B ;
MANY, A .
BRITISH JOURNAL OF HAEMATOLOGY, 1965, 11 (06) :654-&
[25]   Exploiting subsite S1 of trypsin-like serine proteases for selectivity: Potent and selective inhibitors of urokinase-type plasminogen activator [J].
Mackman, RL ;
Katz, BA ;
Breitenbucher, JG ;
Hui, HC ;
Verner, E ;
Luong, C ;
Liu, L ;
Sprengeler, PA .
JOURNAL OF MEDICINAL CHEMISTRY, 2001, 44 (23) :3856-3871
[26]  
MCGRATH ME, 1991, J BIOL CHEM, V266, P6620
[27]   MACROMOLECULAR CHELATION AS AN IMPROVED MECHANISM OF PROTEASE INHIBITION - STRUCTURE OF THE ECOTIN-TRYPSIN COMPLEX [J].
MCGRATH, ME ;
ERPEL, T ;
BYSTROFF, C ;
FLETTERICK, RJ .
EMBO JOURNAL, 1994, 13 (07) :1502-1507
[28]   EXPRESSION OF THE PROTEASE INHIBITOR ECOTIN AND ITS COCRYSTALLIZATION WITH TRYPSIN [J].
MCGRATH, ME ;
ERPEL, T ;
BROWNER, MF ;
FLETTERICK, RJ .
JOURNAL OF MOLECULAR BIOLOGY, 1991, 222 (02) :139-142
[29]   Enhancement of rabbit jugular vein thrombolysis by neutralization of factor XI - In vivo evidence for a role of factor XI as an anti-fibrinolytic factor [J].
Minnema, MC ;
Friederich, PW ;
Levi, M ;
von dem Borne, PAK ;
Mosnier, LO ;
Meijers, JCM ;
Biemond, BJ ;
Hack, CE ;
Bouma, BN ;
ten Cate, H .
JOURNAL OF CLINICAL INVESTIGATION, 1998, 101 (01) :10-14
[30]  
NAITO K, 1991, J BIOL CHEM, V266, P7353