Poster Presentation The 45th Lorne Conference on Protein Structure and Function 2020

Tandem Domain Evasins: Inhibitors of Chemokine-mediated Inflammation (#209)

Ram Prasad Bhusal 1 , Pramod Aryal 1 , Shankar Raj Devkota 1 , Jenni Hayward 1 , Sayeeda Chowdhury 1 , Andrew Perry 2 , Martin Stone 1
  1. Infection and Immunity Program, Monash Biomedicine Discovery Institute, and Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia
  2. Monash Bioinformatics Platform, Monash University, Clayton, VIC 3800, Australia

Inflammation is a complex physiological response to tissue injury or infection. Leukocyte recruitment is a ubiquitous feature of inflammation. Leukocyte migration is regulated by small secreted proteins called chemokines, which bind to G-protein coupled receptors (GPCRs) expressed on target leukocyte subsets [1]. However, in some cases, the response overcompensates, leading to chronic inflammatory diseases. Ticks are hematophagous organisms, which suck blood from their hosts for up to two weeks. As an evolutionary strategy, ticks have been found to secrete glycoproteins called Evasins that bind and block chemokine activity, enabling ticks to avoid detection and prolong blood feeding. As such, Evasins are natural chemokine-neutralising proteins with potential as therapeutics for chronic inflammatory diseases. In recent years, we and others have discovered a large family of tick Evasins that bind and inhibit chemokines [2]. Now by using bioinformatics, we have discovered 82 novel sequences that contain tandem repeats of Evasin-like sequences. It is likely that these “tandem-domain Evasins (TDEs)” can simultaneously recognise two chemokines, potentially in a cooperative manner with functional advantages over the previously characterised Evasins. we have expressed selected TDEs using E. coli and mammalian expression systems. The proteins were purified chromatographically and evaluated by SDS-PAGE, analytical size exclusion chromatography (ASEC), analytical reverse-phase HPLC and mass spectrometry. We have determined the chemokine-binding properties of the selected TDEs by measuring binding to a set of CC and CXC chemokines using a competitive fluorescence anisotropy and surface plasmon resonance assays. Our work may pave the way for the development of new therapeutic agents against inflammation.

 

  1. Bhusal, R. P.; Foster, S. R.; Stone, M. J., Structural basis of chemokine and receptor interactions: Key regulators of leukocyte recruitment in inflammatory responses. Protein Sci 2019.
  2. Bhusal, R. P.; Eaton, J. R. O.; Chowdhury, S. T.; Power, C. A.; Proudfoot, A. E. I.; Stone, M. J.; Bhattacharya, S., Evasins: Tick Salivary Proteins that Inhibit Mammalian Chemokines. Trends Biochem Sci 2019.