Probing interactions between the coagulants thrombin, Factor XIII, and fibrin(ogen).
Thrombin cleaves fibrinopeptides A and B from fibrinogen leading to the formation of a fibrin network that is later covalently crosslinked by Factor XIII (FXIII). Thrombin helps activate FXIII by catalyzing hydrolysis of the FXIII activation peptides (AP). In the current work, the role of exosites in the ternary thrombin-FXIII-fibrin(ogen) complex was further explored. Hydrolysis studies indicate that thrombin predominantly utilizes its active site region to bind extended Factor XIII AP (FXIII AP 33-64 and 28-56) leaving the anion-binding exosites for fibrin(ogen) binding. The presence of fibrin-I leads to improvements in the K(m) for hydrolysis of FXIII AP (28-41), whereas peptides based on the cardioprotective FXIII V34L sequence exhibit less reliance on this cofactor. Surface plasmon resonance measurements reveal that d-Phe-Pro-Arg-chloromethylketone-thrombin binds to fibrinogen faster than to FXIII a(2) and dissociates from fibrinogen more slowly than from FXIII a(2). This system of thrombin exosite interactions with differing affinities promotes efficient clot formation.
Maurer
Muriel C
MC
Department of Chemistry, University of Louisville, 2320 South Brook Street, Louisville, KY 40292, USA. muriel.maurer@louisville.edu
Trumbo
Toni A
TA
Isetti
Giulia
G
Turner
Brian T
BT
D000590:Amino Acid Chloromethyl Ketones / Q000737:chemistry; D000595:Amino Acid Sequence; D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D005176:Factor XIII / Q000737:chemistry*; D005337:Fibrin / Q000737:chemistry*; D005340:Fibrinogen / Q000737:chemistry*; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011485:Protein Binding; D013379:Substrate Specificity; D020349:Surface Plasmon Resonance; D013917:Thrombin / Q000737:chemistry*
D000590:Amino Acid Chloromethyl Ketones; D010446:Peptide Fragments; D005337:Fibrin; D005340:Fibrinogen; D005176:Factor XIII; D013917:Thrombin; C050815:phenylalanyl-prolyl-arginine chloromethyl ketone-thrombin