Physical map location of the Escherichia coli gene encoding acyl coenzyme A synthetase.

E. coli contains a single acyl-CoA synthetase, which has been purified to homogeneity (3, 4). In the process of long-chain fatty acid transport, this enzyme plays a pivotal role by catalyzing the thioesterification of exogenous fatty acids into metabolically active CoA thioesters prior to 3-oxidation concomitant with transport. This enzyme has broad chain-length specificity, giving Vm. values ranging from 2,632 nmollmin/mg of protein for lauric acid (C12) to 135 nmollmin/mg of protein for hexanoate (C6) (3). Maximal activities associated with this enzyme are found with fatty acids ranging in length from C12 to C18:1 (3). The structural gene for acyl-CoA synthetase (fadD) was identified by Overath et al. (6), who mapped this locus to the 40-min region of the E. coli chromosome. I have used 11 clones representing the 40-min region of the E. coli chromosome from the Kohara library (5) (clones 5E12, 4B8, 12H7, 3E12, 9F2, 7F2, 6D1, 12B3, 15D5, 19H3, and 12C7) to fine map and subclone the fadD gene (1). The fadD88 strain PN235 was transduced with these clones along with XCI857 as a helper phage and then plated on oleate minimal agar plates. After 72 h at 30°C, dilysogens that were able to grow on oleate as a sole carbon and energy source (Ole') were identified in cells infected with clones 7F2 and 6D1. A second round of lysogenic complementation demonstrated that clones 7F2 and 6D1 were both able to confer an Ole' phenotype in thefadD88 strain PN235. Clone 6D1 was used as a source of flNA to subclone the fadD gene for further analysis (1). Analysis using restriction endonucleases indicated that thefadD gene is located at approximately 1905 kb, corresponding to 39.5 min on the genetic map (Fig. 1).