Structural basis for broad substrate specificity of earthworm fibrinolytic enzyme component A

• Published in: Biochemical and biophysical research communications 2004-12, Vol.325 (3), p.877-882
• Main Authors: Wang, Chao, Wang, Feng, Li, Mei, Tang, Yong, Zhang, Ji-Ping, Gui, Lu-Lu, An, Xiao-Min, Chang, Wen-Rui
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Binding Sites; Catalysis; Computer Simulation; Earthworm fibrinolytic enzyme; Endopeptidases - analysis; Endopeptidases - chemistry; Enzyme Activation; Models, Chemical; Models, Molecular; Molecular Sequence Data; Multiprotein Complexes - chemistry; Multisubstrate-binding sites; Oligochaeta - enzymology; Protein Binding; Protein Conformation; Protein Structure, Secondary; Structure-Activity Relationship; Substrate Specificity
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2004.10.113

Earthworm fibrinolytic enzyme component A (EFE-a) possesses an S1 1 Nomenclature for the substrate amino acid residues is P n, … , P2, P1, P1′, P2′, … , P n′, where P1–P1′ denotes the hydrolyzed bond. S n, … , S2, S1, S1′, S2′, … , S n′ denotes the corresponding enzyme binding sites. 1 pocket, which is typical for an elastase-like enzyme, but it can still hydrolyze varieties of substrates, and it exhibits wide substrate specificity. Former structure studies suggested that the four-residue insertion after Val 217 2 Chymotrypsinogen numbering is used throughout. 2 might endow EFE-a with this specificity. Based on the native crystal structure at a resolution of 2.3 Å, we improved the native crystal structure to 1.8 Å and determined its complex structure with the inhibitor Meo-Suc-Ala-Ala-Pro-Val-CMK at a resolution of 1.9 Å. The final structures show that: (1) EFE-a possesses multisubstrate-binding sites interacting with the substrates; (2) significant conformation adjustment takes place at two loops binding to the N-terminal of the substrates, which may enhance the interaction between the enzyme and the substrates. These characteristics make the substrate-specificity of EFE-a less dependent on the property of its S1-pocket and may endow the enzyme with the ability to hydrolyze chymotrypsin-specific substrates and even trypsin-specific substrates.