Bypassing the requirement for aminoacyl-tRNA by a cyclodipeptide synthase enzyme

Christopher J. Harding, Emmajay Sutherland, Jane G. Hanna, Douglas R. Houston, Clarissa M. Czekster

Research output: Contribution to journalArticlepeer-review


Cyclodipeptide synthases (CDPSs) produce a variety of cyclic dipeptide products by utilising two aminoacylated tRNA substrates. We sought to investigate the minimal requirements for substrate usage in this class of enzymes as the relationship between CDPSs and their substrates remains elusive. Here, we investigated the Bacillus thermoamylovorans enzyme, BtCDPS, which synthesises cyclo(L-Leu-L-Leu). We systematically tested where specificity arises and, in the process, uncovered small molecules (activated amino esters) that will suffice as substrates, although catalytically poor. We solved the structure of BtCDPS to 1.7 Å and combining crystallography, enzymatic assays and substrate docking experiments propose a model for how the minimal substrates interact with the enzyme. This work is the first report of a CDPS enzyme utilizing a molecule other than aa-tRNA as a substrate, providing insights into substrate requirements and setting the stage for the design of simpler catalytically improved substrates.
Original languageEnglish
Number of pages11
JournalRSC Chemical Biology
Early online date15 Jan 2021
Publication statusE-pub ahead of print - 15 Jan 2021


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