In vivo incorporation of lipoic acid enantiomers and homologues in the pyruvate dehydrogenase complex from Escherichia coli.
The strain Escherichia coli JRG26, which has a defect in the lipoic acid biosynthesis, was cultivated in the presence of R-lipoic acid, S-lipoic acid, RS-dithiolane-3-caproic acid, RS-bisnorlipoic acid, and RS-tetranorlipoic acid, respectively. With the exception of the last compound the strain was able to grow with all these substances. R-lipoic acid was the most efficient factor, concentrations of 10 ng/l were sufficient to support growth of the cells, while 10(4)-fold to 10(7)-fold higher concentrations were necessary for the other compounds. The specific catalytic activity of the pyruvate dehydrogenase complex isolated from the cells grown on RS-dithiolane-3-caproic acid was only slightly lower than from cells grown on R-lipoic acid. With RS bisnorlipoic acid 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 antibodies directed against R-lipoic acid and RS-bisnorlipoic acid, both conjugated to BSA. Western blot analysis showed that the antibodies against the R-lipoic acid reacted specifically with the E2 component of pyruvate dehydrogenase complex purified from cells grown on this factor, while antibodies against RS-bisnorlipoic acid reacted with the enzyme complex isolated from cells grown in the presence of this compound.
Loeffelhardt
S
S
Physiologisch-Chemisches Institut, Universität Tübingen, Germany.
Borbe
H O
HO
Locher
M
M
Bisswanger
H
H
D000123:Acetyltransferases / Q000378:metabolism*; D001665:Binding Sites; D051047:Dihydrolipoyllysine-Residue Acetyltransferase; D004926:Escherichia coli / Q000201:enzymology* / Q000254:growth & development; D011233:Precipitin Tests; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism*; D013237:Stereoisomerism; D008063:Thioctic Acid / Q000031:analogs & derivatives / Q000378:metabolism*
D011768:Pyruvate Dehydrogenase Complex; D008063:Thioctic Acid; D000123:Acetyltransferases; D051047:Dihydrolipoyllysine-Residue Acetyltransferase