Creative Biolabs has established a unique ribosome display platform based on our proprietary prokaryotic and eukaryotic ribosome display technologies. The ribosome display platform is a superior in vitro cell-free platform that generates peptide, protein and antibody libraries with an unparalleled diversity up to 1015; as a result, extremely high-affinity binders against a variety of targets can be isolated from the libraries. In particular, our ribosome display antibody libraries can produce antibodies with pM affinity, the highest affinity ever achieved by an antibody production technology.
Ribosome display is a completely cell-free system for the in vitro selection of proteins and peptides from large libraries. As it is completely performed in vitro, it circumvents many drawbacks of in vivo systems. It can display very large libraries, suitable for generating toxic, proteolytically sensitive and unstable proteins, and also allows amino acids modification at defined position. In ribosome display, an individual nascent protein to its corresponding mRNA couples through the formation of stable protein-ribosome-mRNA (PRM) complexes, and this complex serves as the physical link between genotype and phenotype. mRNAs are isolated from target-binding complexes, reverse transcribed and amplified as DNA for further manipulation and protein expression. The PRM complex is very stable. The integrity of PRM can be maintained for several days under appropriate conditions, which provide a good way for stringent selections. Besides, ribosome display permits the simultaneous isolation of a functional nascent protein, through affinity for a ligand, together with the encoding mRNA.
Figure 1. Principle of ribosome display
Creative Biolabs provides several systems for ribosome display, which include the following:
The first published description of ribosome display was for peptide selection using Escherichia coli S30 extract system. Creative Biolabs provides E. coli S30 cell-free lysate for ribosome display, either with coupled or uncoupled transcription and translation.
Eukaryotic cell-free systems are capable of a variety of posttranslational modifications, expanding the possibility of displaying functionally relevant proteins. Creative Biolabs provides rabbit reticulocyte lysate system for ribosome display. This system employs an in situ recovery procedure in which DNA was recovered directly on the ribosome complexes by amplification with a novel in situ RT-PCR procedure without dissociation of the ribosome complex.
RIDS is perceived to be a powerful technique for the selection and evolution of peptides and proteins, useful for protein engineering. In the case of RIDS, a gene encoding a toxin that inactivates eukaryotic ribosomes, the ricin A chain (RTA) is fused downstream the sequence library to enable the complex formation. As a result, the functional proteins can be isolated under standard translation conditions without the need to remove the termination codon.
Ribosome display technology has many applications, which including but not limited to:
As ribosome display can avoid library limitation and is capable of screening much larger libraries, it is more and more popular in selecting high affinity, specific ligand-binding peptides from combinatorial libraries. We can provide as large as 1012-1014 combinatorial peptide libraries to generate high-quality peptides.
Creative Biolabs has developed ribosome display system to carry out phenotypic selection for scFv against a variety of peptides and proteins targets. During this process, sequence evolution and screening can be performed simultaneously.
Except for screening antibodies, ribosome display can also be used for affinity selection of non-antibody proteins. Creative Biolabs has developed a system that uses affinity tag for isolating PRM-complexes for proteins without being antibodies. The affinity tag can be fused to the N-terminus of the protein for ribosome display. Besides, we can also select peptides/proteins binding to enzymes or display functional proteins for applications such as drug targets identification and protein function identification.
In combination with DNA mutagenesis, ribosome display offers a powerful means of protein evolution in vitro. This approach offers a good way for antibody maturation and improving the biophysical properties. We offer DNA evolution by low-fidelity DNA polymerase, mutagenesis methods, and homologous recombination.
Ribosome display technology is a valuable tool for in vitro directed evolution of protein stability and affinity due to its two major advantages: (1) The diversity of ribosome display libraries is not limited by the transformation efficiency of bacterial cells; (2) Random mutations can be introduced easily into the selected DNA pool in subsequent cycles. Based on this technology, we are confident in offering top-quality end-to-end services for optimizing protein affinity and stability.
Ribosome display, a cell-free display technology, has been applied for in vitro selection of catalytic activity. Combining advanced random mutagenesis technology and ribosome display library construction and screening, Creative Biolabs provides a full range of solutions for the selection and improvement of enzymatic activity.
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