Orphan Three-Finger Toxins Bind at Tissue Factor-Factor VIIa Interface to Inhibit Factor X Activation: Identification of Functional Site by Docking.

Manisha Choudhury,R Manjunatha Kini,Devadasan Velmurugan,Ryan Mccleary

doi:10.1055/s-0038-1672184

Manisha Choudhury, R Manjunatha Kini + Show 2 more

Open Access

https://doi.org/10.1055/s-0038-1672184

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Abstract

Three-finger toxins (3FTxs) contribute to toxicity of venomous snakes belonging to the family Elapidae. Currently, functions of a considerable proportion of 3FTxs are still unknown. Here, we describe the function of orphan group I 3FTxs consisting of four members. We also identified a new member of this group by sequencing a transcript isolated from Naja naja venom. This transcript, named najalexin, is identical to that previously described 3FTx from Naja atra venom gland, and shared high sequence identity with ringhalexin from Hemachatus haemachatus and a hypothetical protein from Ophiophagus hannah (here named as ophiolexin). The three-dimensional structure, as predicted by molecular modeling, showed that najalexin and ophiolexin share the same conserved structural organization as ringhalexin and other 3FTxs. Since ringhalexin inhibits the activation of factor X by the tissue factor–factor VIIa complex (TF-FVIIa), we evaluated the interaction of this group of 3FTxs with all components using in silico protein–protein docking studies. The binding of orphan group I 3FTxs to TF-FVIIa complex appears to be driven by their interaction with TF. They bind to fibronectin domain closer to the 170-loop of the FVIIa heavy chain to inhibit factor X activation. The docking studies reveal that functional site residues Tyr7, Lys9, Glu12, Lys26, Arg34, Leu35, Arg40, Val55, Asp56, Cys57, Cys58, and Arg65 on these 3FTxs are crucial for interaction. In silico replacement of these residues by Ala resulted in significant effects in the binding energies. Furthermore, these functional residues are not found in other groups of 3FTxs, which exhibit distinct pharmacological properties.

Highlights

Since ringhalexin inhibits the activation of factor X by the tissue factor–factor VIIa complex (TF-FVIIa), we evaluated the interaction of this group of 3FTxs with all components using in silico protein–protein docking studies
Snake venom is an adaptive evolutionary innovation that consists of a mixture of proteins and polypeptides, a large number of which exhibit diverse biological activities
To understand the interaction of ringhalexin and other orphan group I 3FTxs with coagulation factors involved in the extrinsic tenase complex and to determine structure– function relationships of these toxins, we used molecular docking experiments. To this end, we modeled the structures of najalexin and ophiolexin based on the crystal structure of ringhalexin (PDB code 4ZQY)[27] and studied the interaction of these proteins with FVIIa, TF, FX, and TF-FVIIa using in silico protein–protein docking approaches

Summary

Introduction

Snake venom is an adaptive evolutionary innovation that consists of a mixture of proteins and polypeptides, a large number of which exhibit diverse biological activities. TF Drives the Binding to Extrinsic Tenase Complex The toxins docked with FVIIa and FX showed interactions at the interface of the heavy (H) and light (L) chains but on the opposite surface of their active sites (►Fig. 4A, B; see ►Supplementary Fig. S4A, B, E, F; ►Table 1). Najalexin appears to interact with TF using four residues Tyr[54], Arg[60], Cys[63], and Arg[65]

Results

Conclusion