Biosynthesis of lipoic acid: Characterization of the lipoic acid auxotrophs Escherichia coli W1485-lip2 and JRG33-lip9

Hayden, M.A.; Huang, I.Y.; Iliopoulos, G.; Orozco, M.; Ashley, G.W.

Biochemistry 32(14): 3778-3782


ISSN/ISBN: 0006-2960
PMID: 8466915
DOI: 10.1021/bi00065a033
Accession: 008233508

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The abilities of the Escherichia coli lipoic acid auxotrophs W1485-lip2 and JRG33-lip9 to grow on succinate medium in the presence of octanoate, 8-mercaptooctanoate, or 6-mercaptooctanoate have been determined. Both organisms are mutated in lipA. Neither organism can use octanoate or 6-mercaptooctanoate for production of lipoate, but the lip2 allele can use 8-mercaptooctanoate. Chromosomal DNA from the auxotrophs was amplified by PCR using primers derived from the DNA sequence of wild-type lipA and then sequenced. Both mutants contain single G/C to A/T mutations in lipA, resulting in conversion of Ser-307 into Phe in W1485-lip2 and Glu-195 into Lys in JRG33-lip9. These results support the hypothesis that lipA is involved in the sulfur insertion step(s) of lipoate biosynthesis and indicate that it is possible to selectively block formation of the C-8 sbd S bond through suitable mutation in lipA.