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EETI-II Library Construction

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Creative Biolabs provides unique custom EETI-II scaffold library construction service with satisfactory controllability and operability. Our Hi-Affi™ EETI-II library with high quality and diversity is suitable for screening and selecting of high affinity target-binders.

The Ecballium elaterium trypsin inhibitor II (EETI-II), isolated from squirting cucumber seeds, is a kind of squash inhibitors, which is one of the strongest inhibitors known for trypsin. EETI-II is a 28 amino acids protein belonging to the knottin family (also known as microbody or miniprotein). This family represents a group of very small proteins containing six conserved cysteine residues shaping the cystine-stabilized β-sheet (CSB) motif, which is composed of a cystine-knot that arises from three interlocking disulfide bridges and a small triple stranded antiparallel β-sheet. The family members also present an inserted peptide loop with high variability in both sequence and length. The tolerance of this exposed loop has been demonstrated for the integration of defined binding domains. Due to the similar architecture among knottin family members, EETI-II is sufficiently rigid and stable, which can efficiently configure all of its disulfide bonds and correctly fold in vitro, which facilitates its chemical synthesis. According to the structure analysis and remarkable stability, the exposed loop of EETI-II is considered variable in sequence but stable in conformation. Therefore, the variants of exposed loop can enable EETI-II to be qualified as a molecular scaffold for protein engineering purposes. Through the advanced phage display technology of Creative Biolabs, high affinity and diversity EETI-II libraries can be generated for different application.

Phage display is an exogenous gene expression method that the recombinant peptides or proteins are fused with a coated protein of bacteriophage and thereby displayed on the phage surface. It has become a powerful method for screening and isolating proteins that have novel-binding activities. Creative Biolabs has developed a proprietary Hi-Affi™ phage display platform based on trimer codon technology and NNK method, which can create a large repertoire of variants for specific protein scaffolds with 100% precise mutant and over 1010 diversity.

With extensive experience in the field of scaffold library construction, Creative Biolabs has generated about 55 different scaffold libraries to meet our global clients’ requirements. Our scientists are confident in offering the best service to accelerate customers’ research project with high efficiency and reasonable price.

Crystal structure of the free (uncomplexed) Ecballium elaterium trypsin inhibitor (EETI-II) Fig.1 Crystal structure of the free (uncomplexed) Ecballium elaterium trypsin inhibitor (EETI-II). (PDB ID: 1W7Z)




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