Biomedical Engineering Reference
In-Depth Information
FIGURE 11.1
Screening vs. selection approaches. (a) Hundreds of thousands to millions
of chemical compounds are discretely “screened” against a target protein. Only proteins for
which a biochemical assay has been developed can be used in screening campaigns. Such an
approach (i.e., high-throughput screening) is extremely demanding in terms of costs, time, and
logistics. (b) Very large repertoires of biomacromolecules can be selected simultaneously by
affinity capture against virtually any target protein of choice. After applying selection pressure,
nonbinders are washed away, while binding compounds are amplified by bacterial infection
and identified by DNA sequencing of the corresponding “genotype.” Eventually, to improve
the binding properties of the enriched candidates, a second-generation (affinity maturation)
library can be reconstituted from the selected population and serve as an input for further
rounds of selection. The entire process can be performed in just a few days.
In sharp contrast to the isolation of small organic molecules, the identification
of specific biopolymer ligands to virtually any biological target antigen can be
successfully and elegantly accomplished by applying display techniques such as
phage display [26-29], yeast display [30], ribosome display [31,32],
m
RNA dis-
play [33-35], covalent display [36], and other conceptually similar methodologies
[37,38]. Different from high-throughput screening, such technologies rely on the
rapid assembling and selection of large repertoires of functional polypeptides (e.g.,
antibody libraries) containing billions of library members [39]. By means of native
biosynthetic machineries, highly complex mixtures of recombinant DNA-genotype
are individually translated into the corresponding molecules (i.e., proteins, RNA).
Subsequently, after imposing a selection pressure on the entire library of candidates,
compounds with optimal binding properties are preferentially enriched. The intimate
linkage between the displayed phenotype (e.g., antibody) and the corresponding
