The sgaTBA genes of Escherichia coli encode a putative 12-transmembrane ␣-helical segment (12 TMS) transporter, an enzyme IIB-like protein and an enzyme IIA-like protein of the phosphotransferase system (PTS), respectively. We show that all three proteins as well as the energy-coupling PTS proteins, enzyme I and HPr, are required for the anaerobic utilization and uptake of L-ascorbate in vivo and its phosphoenolpyruvatedependent phosphorylation in vitro. The transporter exhibits an apparent K m for L-ascorbate of 9 M and is highly specific. The sgaTBA genes are regulated at the transcriptional level by the yjfQ gene product, as well as by Crp and Fnr. The yjfR gene product is essential for L-ascorbate utilization and probably encodes a cytoplasmic L-ascorbate 6-phosphate lactonase. We conclude that SgaT represents a novel prototypical enzyme IIC that functions with SgaA and SgaB to allow phosphoryl transfer from HPr(his-P) to L-ascorbate via the phosphoryl transfer pathway: PEP ¡ enzyme I-P ¡ HPr-P ¡ IIA-PIn 1996, we reported computational analyses of three operons present in the Escherichia coli genome that appeared to encode enzymes and a transporter concerned with sugar metabolism (18). These operons, sga, sgb, and sgc, encode enzymes homologous to pentose-phosphate 3-and 4-epimerases, and sga and sgc also encode homologues of constituents of the bacterial phosphotransferase system (PTS) (22). The sga operon includes two genes, sgaA and sgaB, that encode homologues of fructose/mannitol enzymes IIA and lactose/N,NЈ-diacetylchitobiose enzymes IIB, respectively ( Fig. 1). No IIC homologue was identified, but upstream of the sgaB and sgaA genes is a gene, sgaT, that encodes a membrane protein with 12 putative transmembrane helical segments, a characteristic of many secondary carriers (25). SgaT proved not to be homologous to any functionally characterized protein. Two possibilities were considered: (i) SgaT-SgaB-SgaA could be a novel type of enzyme II complex, or (ii) SgaT could be a secondary transporter, while SgaA and SgaB might regulate expression of the sga operon or the activity of one or more of its gene products (18,25).It has been known for over 60 years that various bacteria, including E. coli, can ferment L-ascorbate (L-xyloascorbate [vitamin C]) under anaerobic but not aerobic conditions (7,19,32). The unstable hydrolysis product of L-ascorbate, 3-keto-Lgulonate, has been implicated as an intermediate in its catabolism (28). The dissimilation of L-ascorbate, both during coculture with other carbon sources and as a sole carbon source, has been documented for E. coli (19,31). In animal tissues, the principal route of L-ascorbate metabolism involves enzymatic and nonenzymatic oxidation to dehydroascorbate, an unstable lactone that hydrolyzes spontaneously to 2,3-di-keto-L-gulonate (9).Recently, Yew and Gerlt (31) showed that the anaerobic utilization of L-ascorbate is dependent on three enzymes encoded downstream of sgaTBA within the sga operon (Fig. 1). These enzymes catalyze the conversion of the phosp...