The cai E gene, which encodes an enzyme involved in the synthesis or the activation of the still unknown cofactor required for carnitine dehydratase and carnitine recemase activities, was first iso lated by PCR from a genomic library of E.coli K12s, then was cloned into cloning vector pBluescript SK. Sequence analysis shows that the cai E gene is 612 bp long, with 10 nucletides and 6 deduced amino acids different from that reported. The gene was ligated into different vectors to construct expression vectors with different duplicons and different promoters pET CaiE, pWSK CaiE and pACYC CaiE. Then these expression vectors were transformed into E.coli BL21(DE3), after induction with 1mmol/L IPTG, CaiE was highly expressed. SDS PAGE showed the molecular weight of CaiE was 26ku, and the proportion of expression product in the total bacteria protein was pET CaiE 30%, pACYC CaiE 8%, pSC CaiE 5%, respectively.