Abstract
Assembly line enzymes, including polyketide synthases and nonribosomal peptide synthetases, play central roles in the construction of complex natural products. Due to the sequential biochemistry processed in each domain, the domain architecture of the assembly line enzymes strictly correlates with the product molecule. This colinearity makes assembly line enzymes an ideal target for rational reprogramming. Although many of the past engineering attempts suffered from decreased product yield, recent advancements in the bioinformatic analysis and engineering design now provide new opportunity to work on these modular megaenzymes. This chapter describes the methods for analyzing and engineering the assembly line enzymes, including module and domain analysis needed for designing the engineering of assembly line biosynthesis, and the expression vector construction with an example of two-vector heterologous expression system in Streptomyces.
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