Inhibition of PD1:PD-L1 interaction by an E. coli-derived optimized PD1 variant

Michal Brand Shwartz, Mayan Assor, Nesly Dotan, Einav Ratzon, Elad Cohen, Nili Ruimi, Itai Bloch, Maayan Gal, Itamar Yadid

Research output: Contribution to journalArticlepeer-review

Abstract

Immune-checkpoint receptors are a set of signal transduction proteins that can stimulate or inhibit specific anti-tumor responses. It is well established that cancer cells interact with different immune checkpoints to shut down T-cell response, thereby enabling cancer proliferation. Given the importance of immune checkpoint receptors, a structure-function analysis of these systems is imperative. However, recombinant expression and purification of these membrane originated proteins is still a challenge. Therefore, many attempts are being made to improve their expression and solubility while preserving their biological relevance. For this purpose, we designed an E. coli-based optimization system that enables the acquisition of mutations that increases protein solubility and affinity towards its native ligand, while maintaining biological activity. Here we focused on the well-characterized extracellular domain of the 'programmed cell death protein 1' (PD1), an immune checkpoint receptor known to inhibit T-cell proliferation by interacting with its ligands PD-L1 and PD-L2. The simple ELISA-based screening system shown here enabled the identification of high-affinity, highly soluble, functional variants derived from the extracellular domain of human PD1. The system was based on the expression of a GST-tagged variants library in E. coli, which enabled the selection of improved PD1 variants after a single optimization round. Within only two screening rounds, the most active variant showed a 5-fold higher affinity and 2.4-fold enhanced cellular activity as compared to the wild type protein. This scheme can be translated toward other types of challenging receptors toward development of research tools or alternative therapeutics.

Original languageEnglish
Pages (from-to)731-738
Number of pages8
JournalBiochemical and Biophysical Research Communications
Volume506
Issue number3
DOIs
StatePublished - 30 Nov 2018
Externally publishedYes

Keywords

  • Directed evolution
  • E. coli-based ELISA
  • PD1:PD-L1 interaction
  • Protein optimization
  • Soluble receptors

Fingerprint

Dive into the research topics of 'Inhibition of PD1:PD-L1 interaction by an E. coli-derived optimized PD1 variant'. Together they form a unique fingerprint.

Cite this