UC Berkeley

The soil-dwelling bacterium Streptomyces cattleya produces the antibiotics fluoroacetate and fluorothreonine, and has served as a model system for the discovery of naturally occurring fluorine-selective biochemistry. While fluoroacetate has long been known to act as an inhibitor of the TCA cycle, the fate of the amino acid fluorothreonine is still not well understood. Here, I show that while fluorothreonine is a substrate for translation, this activity is averted in S. cattleya by the activity of two conserved proteins. The first, SCAT_p0564, acts in vitro and in vivo as a fluorothreonyl-tRNA selective hydrolase, while the second, SCAT_p0565, is proposed to be a fluorothreonine exporter. Additionally, overexpression of SCAT_p0564 in the model strain Streptomyces coelicolor M1152 confers resistance to fluorothreonine, suggesting that the antibiotic activity of this compound is related in part to its ability to enter the proteome. The ability of SCAT_p0564 to selectively hydrolyze fluorothreonyl- over threonyl-tRNA is striking, given that these macromolecular substrates differ by a single atom. In order to understand the basis of this selectivity, I have solved the crystal structure of this enzyme, and characterized its ability to act on the related substrate chlorothreonyl-tRNA. I also have also begun to elucidate the regulatory architecture of the organofluorine biosynthesis locus, with the aim of understanding how this unusual process is controlled. I find that transcription of the fluorinase is driven by the master regulator FlG, while FlF is an amino acid binding transcription factor that may be required for full expression of the fluorothreonine transaldolase. These findings expand the range of known naturally occurring fluorine biochemistry, and a represent a step towards the rational discovery of new organofluorine metabolism in nature.