The optimization of a metabolic pathway is a time-consuming process, dominated by multiple rounds of trial-and-error mutation, with the goal of finding the optimal enzyme expression levels that maximizes a product yield. To greatly accelerate this process, we have developed a predictive design method that allows one to rationally select the expression level of a protein in bacteria. The method designs the sequence of a synthetic ribosome binding site to achieve a user-selected protein expression level. It correctly predicts the expression level of a designed sequence to within a factor of 2 with a range of over 100,000 expression. As a test example, we apply the design method to optimize a seven enzyme metabolic pathway that synthesizes a carotenoid photopigment.

With the emergence of low cost DNA synthesis and the rapid assembly of genetic and metabolic systems, the development bottleneck now shifts towards identifying the optimal DNA sequence that yields a high performance system. These and other predictive design methods provide that much-needed missing link between sequence and function.

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