Unlike conventional metabolic pathways that lead to a single product, diversity-generating pathways produce many compounds. Understanding such diversity-generating pathways could help researchers develop combinatorial biosynthetic pathways. A team of researchers led by Baldomero M. Olivera and Eric W. Schmidt of the University of Utah shows how one such pathway from cyanobacteria differs significantly from conventional metabolic pathways. The researchers focused on the tru pathway, which synthesizes a diverse family of macrocyclic peptides called patellins (Proc. Natl. Acad. Sci. USA 2016, DOI: 10.1073/pnas.1525438113). To study the four-enzyme pathway, the researchers expressed it in Escherichia coli bacteria. They found that they could greatly increase the yield of patellins by adding cysteine, which releases hydrogen sulfide that modulates the enzymes’ activity. In the process of figuring out the cysteine effect, they learned that each step in the pathway is much slower than the preceding steps. This results in the accumulation of intermediates, which is in sharp contrast to conventional pathways, in which the first step is usually the rate-limiting step to avoid such accumulation.