Biosynthesis of lipoic acid: characterization of the lipoic acid auxotrophs Escherichia coli W1485-lip2 and JRG33-lip9.
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 Ser307 into Phe in W1485-lip2 and Glu195 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 C8-S bond through suitable mutation in lipA.
Hayden
M A
MA
Department of Chemistry and of Biochemistry, Northwestern University, Evanston, Illinois 60208.
Huang
I Y
IY
Iliopoulos
G
G
Orozco
M
M
Ashley
G W
GW
D000595:Amino Acid Sequence; D001483:Base Sequence; D002210:Caprylates / Q000378:metabolism; D004269:DNA, Bacterial / Q000737:chemistry; D015247:Deoxyribonuclease HindIII / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development*; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction; D008063:Thioctic Acid / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics
D002210:Caprylates; D004269:DNA, Bacterial; D008063:Thioctic Acid; D015247:Deoxyribonuclease HindIII; C031492:octanoic acid