Localization of Target-Site of the Protoporphyrinogen Oxidase-Inhibiting Herbicide, S-23142, in Spinacia oleracea L.

Abstract

Effects of S-23142 on protoporphyrin IX (Proto IX) biosynthesis in chloroplasts isolated from Spinacia oleracea L. were examined using reverse phase HPLC with fluorescence monitoring. The synthesis of Proto IX was inhibited to a level of 50% by 10-9 ᴍ of S-23142 in this system. The effects of S-23142 was also tested in chloroplasts isolated from two types of photomixotrophic tobacco cells, wild type and S-23142 tolerant cells. The biosynthesis of both the wild type cells and YZI-1 S cells were inhibited at 50% by 10-9 ᴍ and 10-7 ᴍ of S-23142, respectively. It is, therefore evident that the mutation in the tolerant cell is associated with the Protox. To investigate the localization of Proto IX biosynthesis and the target site of S-23142, spinach chloroplasts were osmotically broken and separated into stroma and membrane (thylakoid and envelope) fractions. A very active Proto IX synthesis from ALA was found in the stromal fraction, while no activity of Proto IX synthesis was observed in the membrane fraction. These results suggest that most Proto IX synthetic activity and a target-site of S-23142 exist in the stromal fraction.