IN-VIVO INCORPORATION OF LIPOIC ACID ENANTIOMERS AND HOMOLOGS IN THE PYRUVATE-DEHYDROGENASE COMPLEX FROM ESCHERICHIA-COLI

The strain Escherichia coli JRG26, which has a defect in the lipoic ac id biosynthesis, was cultivated in the presence of R-lipoic acid, S-li poic acid, RS-dithiolane-3-caproic acid, RS-bisnorlipoic acid, and RS- tetranorlipoic acid, respectively. With the exception of the last comp ound the strain was able to grow with all these substances. R-lipoic a cid was the most efficient factor, concentrations of 10 ng/l were suff icient to support growth of the cells, while 10(4)-fold to 10(7)-fold higher concentrations were necessary for the other compounds. The spec ific catalytic activity of the pyruvate dehydrogenase complex isolated from the cells grown on RS-dithiolane-3-caproic acid was only slighly lower than from cells grown on R-lipoic acid. With RS-bisnorlipoic ac id the specific activity was one third compared to that of the native enzyme complex. The incorporation of the RS-bisnorlipoic acid into the pyruvate dehydrogenase could directly be demonstrated by polyclonal a ntibodies directed against R-lipoic acid and RS-bisnorlipoic acid, bot h conjugated to BSA. Western blot analysis showed that the antibodies against the R-lipoic acid reacted specifically with the E2 component o f pyruvate dehydrogenase complex purified from cells grown on this fac tor, while antibodies against RS-bisnorlipoic acid reacted with the en zyme complex isolated from cells grown in the presence of this compoun d.