, 1999; Degrassi et al., 2002). The results suggest that dipeptide-like compounds such as diketopiperazines are ubiquitous in a natural environment as antimicrobial agents or cell–cell communication molecules (Holden et al., 2000). A blast search revealed that the homologs of Bcr, NorE, YdeE and YeeO prevail in Enterobacteriaceae (data not shown). It was reported that cyclo (Ala-Val) was found in cell-free supernatants from Proteus,

Citrobacter and Enterobacter (Holden et al., 1999). Interestingly, these genera possess close homologs of dipeptide transporters. Taken together, dipeptide transporters and their homologs could be involved in the transport of diketopiperazines in these bacteria. Although it is too early to speculate on High Content Screening the natural functions, these multidrug-efflux transporters may transport dipeptide-like molecules to extracellular space to maintain homeostasis, or as signals for cell–cell communications. It was recently reported that the natural function of NorE was to export signals for cell–cell communication (Yang et al., 2006). In this study, two functionally uncharacterized genes, ydeE and yeeO, were identified as those conferring dipeptide

resistance. Although the minimum inhibitory selleck compound library concentration (MIC) of various antimicrobial agents and chemical compounds were examined in E. coli cells overexpressing ydeE (ydeF), no alteration of MIC was observed (Nishino & Yamaguchi, 2001). In Erwinia chrysanthemi, it is reported that yeeO is a member of the predicted regulon of KdgR, the transcriptional regulator of pectin catabolism O-methylated flavonoid (Rodionov et al., 2004). However, no other information about its function is obtained. In contrast, the effects of ydeE or yeeO overexpression on the reduction of intracellular Ala-Gln and also on the production of Ala-Gln and Ala-BCAA were remarkable (Figs 3 and 4). Bcr and YdeE are classified into the major facilitator superfamily. As shown in Table 2, E. coli cells overexpressing Bcr were resistant to bicyclomycin, tetracycline, fosfomycin, kanamycin and l-cysteine.

NorE and YeeO are classified into the multidrug and toxic compound extrusion family. NorE was identified as a quinolone resistance protein at first (Morita et al., 1998). Also, E. coli cells overexpressing NorE were resistant to chloramphenicol, doxorubicin, fosfomycin, trimethoprim, etc. (Nishino & Yamaguchi, 2001). Although the structure of known substrates is rather dissimilar to Ala-Gln or Ala-BCAA, significant effects on dipeptides production suggest that Bcr, NorE, YdeE and YeeO are involved in the transport of dipeptides. Substrate specificities of these proteins remain to be elucidated. The spectrums of dipeptide to which dipeptide transporters conferred resistance were wide and also differed correspondingly (Table S1).