Identifying the Metal That Activates the Prenyltransferase That Catalyzes Formation of Geranyl Diphosphate in the Diatom Pseudo‐nitzschia multiseries

Abstract

Domoic acid (DA) is a neurotoxin synthesized by marine diatoms of the genus Pseudo‐nitzschia which causes amnesiac shellfish poisoning in humans and death of marine wildlife. Geranyl diphosphate (GPP), a key intermediate in domoic acid biosynthesis, is formed from the “head‐to‐tail” condensation of isopentenyl diphosphate and dimethylallyl diphosphate in a reaction catalyzed by a member of the prenyltransferase family of enzymes. These prenyltransferases can generate different chain‐length products, including geranyl diphosphate (C10), farnesyl diphosphate (FPP, C15), and geranylgeranyl diphosphate (GGPP, C20), all of which are precursors to terpenoid natural products. Prenyltransferases require a divalent metal cation for activity. Previous work has shown that a single prenyltransferase can generate multiple products (GPP, FPP, and GGPP), and that the product distribution will vary depending on the species of divalent cation used to support the reaction (Mg2+, Mn2+, or Co2+). Here, we measured the levels of Mg2+, Mn2+, and Co2+ in Pseudo‐nitzschia cells to determine which ion is most likely to support GPP formation. Three samples of Pseudo‐nitzschia multiseries cultured in sea water were harvested by centrifugation. Samples were digested with HNO3 before analysis using atomic absorption spectroscopy. The average concentration of Mg2+ present, per cell and per μm3, is over 100‐fold higher than the concentrations of Mn2+ and Co2+. This is consistent with results from studies in other systems, suggesting that Mg2+ is the main cofactor that activates the prenyltransferase that generates GPP for domoic acid biosynthesis in Pseudo‐nitzschia.Support or Funding InformationThis project is supported by the SURE (Summer Undergraduate Research Experience) Award Program from the College of Natural Science and Mathematics, California State University Sacramento.