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Biochimie. 2016 Sep-Oct;128-129:138-47. doi: 10.1016/j.biochi.2016.08.005. Epub 2016 Aug 12.

Structural and functional characterization of complex formation between two Kunitz-type serine protease inhibitors from Russell's Viper venom.

Author information

1
Microbial Biotechnology and Protein Research Laboratory, Department of Molecular Biology and Biotechnology, Tezpur University, Tezpur 784028, Assam, India; School of Biological Sciences, University of Northern Colorado, Greeley, CO 80639-0017, USA. Electronic address: akm@tezu.ernet.in.
2
Microbial Biotechnology and Protein Research Laboratory, Department of Molecular Biology and Biotechnology, Tezpur University, Tezpur 784028, Assam, India.
3
Advanced Functional Material Laboratory, Department of Physics, Tezpur University, Tezpur 784028, Assam, India.
4
School of Biological Sciences, University of Northern Colorado, Greeley, CO 80639-0017, USA.

Abstract

Snake venom Kunitz-type serine protease inhibitors (KSPIs) exhibit various biological functions including anticoagulant activity. This study elucidates the occurrence and subunit stoichiometry of a putative complex formed between two KSPIs (Rusvikunin and Rusvikunin-II) purified from the native Rusvikunin complex of Pakistan Russell's Viper (Daboia russelii russelii) venom (RVV). The protein components of the Rusvikunin complex were identified by LC-MS/MS analysis. The non-covalent interaction between two major components of the complex (Rusvikunin and Rusvikunin-II) at 1:2 stoichiometric ratio to form a stable complex was demonstrated by biophysical techniques such as spectrofluorometric, classical gel-filtration, equilibrium gel-filtration, circular dichroism (CD), dynamic light scattering (DLS), RP-HPLC and SDS-PAGE analyses. CD measurement showed that interaction between Rusvikunin and Rusvikunin-II did not change their overall secondary structure; however, the protein complex exhibited enhanced hydrodynamic diameter and anticoagulant activity as compared to the individual components of the complex. This study may lay the foundation for understanding the basis of protein complexes in snake venoms and their role in pathophysiology of snakebite.

KEYWORDS:

Kunitz-type serine protease inhibitor; Phospholipase A(2); Protease; Protein-protein interaction; Snake venom complex

PMID:
27523780
DOI:
10.1016/j.biochi.2016.08.005
[Indexed for MEDLINE]

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