Products/Services Used | Details | Operation |
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Mutagenesis Services> | Site-saturation mutagenesis (SSM) libraries were constructed from synthetic DNA from GenScript. Combinatorial libraries were constructed from synthetic DNA from GenScript containing ambiguous nucleotides and similarly transformed into a linearized pETcon3 vector. Genes encoding the designed protein sequences were synthesized and cloned into pET-28b(+) E. coli plasmid expression vectors (GenScript, N-terminal 6xHis-tagged followed by a thrombin cleavage site. | Get A Quote |
We describe a de novo computational approach for designing proteins that recapitulate the binding sites of natural cytokines, but are otherwise unrelated in topology or amino acid sequence. We use this strategy to design mimics of the central immune cytokine interleukin-2 (IL-2) that bind to the IL-2 receptor βγ heterodimer (IL-2Rβγ) but have no binding site for IL-2Rα (also called CD25) or IL-15Rα (also known as CD215). The designs are hyper-stable, bind human and mouse IL-2Rβγ with higher affinity than the natural cytokines, and elicit downstream cell signalling independently of IL-2Rα and IL-15Rα. Crystal structures of the optimized design neoleukin-2/15 (Neo-2/15), both alone and in comple... More