title: A new peptide (Ruviprase) purified from the venom of Daboia russelii russelii shows potent anticoagulant activity via non-enzymatic inhibition of thrombin and factor Xa.
abstract: Compounds showing dual inhibition of thrombin and factor Xa (FXa) are the subject of great interest owing to their broader specificity for effective anticoagulation therapy against cardiovascular disorders. This is the first report on the functional characterization and assessment of therapeutic potential of a 4423.6 Da inhibitory peptide (Ruviprase) purified from Daboia russelii russelii venom. The secondary structure of Ruviprase is composed of α-helices (61.9%) and random coils (38.1%). The partial N-terminal sequence (E(1)-V(2)-X(3)-W(4)-W(5)-W(6)-A(7)-Q(8)-L(9)-S(10)) of Ruviprase demonstrated significant similarity (80.0%) with an internal sequence of apoptosis-stimulating protein reported from the venom of Ophiophagus hannah and Python bivittatus; albeit Ruviprase did not show sequence similarity with existing thrombin/FXa inhibitors, suggesting its uniqueness. Ruviprase demonstrated a potent in vitro anticoagulant property and inhibited both thrombin and FXa following slow binding kinetics. Ruviprase inhibited thrombin by binding to its active site via an uncompetitive mechanism with a Ki value and dissociation constant (KD) of 0.42 μM and 0.46 μM, respectively. Conversely, Ruviprase demonstrated mixed inhibition (Ki = 0.16 μM) of FXa towards its physiological substrate prothrombin. Furthermore, the biological properties of Ruviprase could not be neutralized by commercial polyvalent or monovalent antivenom. Ruviprase at a dose of 2.0 mg/kg was non-toxic and showed potent in vivo anticoagulant activity after 6 h of intraperitoneal treatment in mice. Because of the potent anticoagulant property as well as non-toxic nature of Ruviprase, the possible application of the peptide as an antithrombotic agent for combating thrombosis-associated ailments appears promising.
authors.affiliation: Microbial Biotechnology and Protein Research Laboratory, Department of Molecular Biology and Biotechnology, School of Science, Tezpur University, Tezpur 784 028, Assam, India.
authors.forename: Rupamoni
authors.initials: R
authors.lastname: Thakur
authors.affiliation: Department of Biotechnology, Indian Institute of Technology, Guwahati 781 039, Assam, India.
authors.forename: Ashok
authors.initials: A
authors.lastname: Kumar
authors.affiliation: Department of Biotechnology, Indian Institute of Technology, Guwahati 781 039, Assam, India.
authors.forename: Biplab
authors.initials: B
authors.lastname: Bose
authors.affiliation: Department of Biosciences and Bioengineering, Indian Institute of Technology, Mumbai 400 076, Maharashtra, India.
authors.forename: Dulal
authors.initials: D
authors.lastname: Panda
authors.affiliation: Microbial Biotechnology and Protein Research Laboratory, Department of Molecular Biology and Biotechnology, School of Science, Tezpur University, Tezpur 784 028, Assam, India.
authors.forename: Debashree
authors.initials: D
authors.lastname: Saikia
authors.affiliation: Division of Pharmaceutical Technology, Defense Research Laboratory, Tezpur 784 001, Assam, India.
authors.forename: Pronobesh
authors.initials: P
authors.lastname: Chattopadhyay
authors.affiliation: Microbial Biotechnology and Protein Research Laboratory, Department of Molecular Biology and Biotechnology, School of Science, Tezpur University, Tezpur 784 028, Assam, India. Electronic address: akm@tezu.ernet.in.
authors.forename: Ashis K
authors.initials: AK
authors.lastname: Mukherjee
mesh_terms: D000818:Animals; D000925:Anticoagulants / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D015374:Biosensing Techniques; D015951:Factor Xa / Q000737:chemistry / Q000378:metabolism; D065427:Factor Xa Inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D051379:Mice; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D060066:Protein Interaction Maps; D012910:Snake Venoms / Q000737:chemistry*; D013917:Thrombin / Q000037:antagonists & inhibitors; D017819:Viperidae
chemical_list: D000925:Anticoagulants; D065427:Factor Xa Inhibitors; D010455:Peptides; D012910:Snake Venoms; D013917:Thrombin; D015951:Factor Xa
keywords: Anticoagulant peptide; Biosensor analysis; Daboia russelii russelii; Factor Xa inhibitor; Protein–protein interaction; Thrombin inhibitor