Differential expression of a putative CarD-like transcriptional regulator, LtpA, in Borrelia burgdorferi.

The availability of microbial genome information has provided a fruitful opportunity for studying regulatory networks in a variety of pathogenic bacteria. In an initial effort to elucidate regulatory networks potentially involved in differential gene expression by the Lyme disease pathogen Borrelia burgdorferi, we have been investigating the functions and regulation of putative transcriptional regulatory factors predicted to be encoded within the B. burgdorferi genome. Herein we report the regulation of one of the predicted transcriptional regulators, LtpA (BB0355), which is homologous to the transcriptional regulator CarD from Myxococcus xanthus. LtpA expression was assessed in response to various environmental stimuli. Immunoblot and quantitative reverse transcription-PCR analyses revealed that unlike many well-characterized differentially regulated Borrelia genes whose expression is induced by elevated temperature, the expression of LtpA was significantly downregulated when spirochetes were grown at an elevated temperature (37 degrees C), as well as when the bacteria were cultivated in a mammalian host-adapted environment. In contrast, LtpA was induced at a lower culture temperature (23 degrees C). Further analyses indicated that the downregulation of LtpA was not dependent on the Rrp2-RpoN-RpoS regulatory pathway, which is involved in the downregulation of OspA when B. burgdorferi is grown in a mammalian host-adapted environment. LtpA protein levels in B. burgdorferi were unaltered in response to changes in the pH in the borrelial cultures. Multiple attempts to generate an LtpA-deficient mutant were unsuccessful, which has hampered the elucidation of its role in pathogenesis. Given that LtpA is exclusively expressed during borrelial cultivation at a lower temperature, a parameter that has been widely used as a surrogate condition to mimic B. burgdorferi in unfed (flat) ticks, and because LtpA is homologous to a known transcriptional regulator, we postulate that LtpA functions as a regulator modulating the expression of genes important to B. burgdorferi's survival within its arthropod vector.

The Tp38 (TpMglB-2) lipoprotein binds glucose in a manner consistent with receptor function in Treponema pallidum.

A 38-kDa lipoprotein of Treponema pallidum (Tp38) was predicted to be a periplasmic sugar-binding protein based on its sequence similarity to the glucose/galactose-binding (MglB) protein of Escherichia coli (P. S. Becker, D. R. Akins, J. D. Radolf, and M. V. Norgard, Infect. Immun. 62:1381-1391, 1994). Inasmuch as glucose is believed to be the principal, if not sole, carbon and energy source for T. pallidum and is readily available to the spirochete during its obligate infection of humans, we hypothesized that Tp38 may serve as the organism's requisite glucose receptor. For the present study, a nonacylated recombinant form of Tp38 was coexpressed with GroES and GroEL in E. coli to facilitate the isolation of soluble, properly folded Tp38. The highly sensitive method of intrinsic fluorescence spectroscopy, predicated on the manner in which tryptophan residues reside and move within protein microenvironments, was then used to assess sugar binding to Tp38. The intrinsic fluorescence of Tp38 was essentially unaltered when it was exposed to D-mannose, D-fucose, D-ribose, L-glucose, or L-galactose, but it changed markedly in the presence of D-glucose, and to a lesser extent, D-galactose, indicating binding. The K(d) values for D-glucose and D-galactose binding to Tp38 were 152.2 +/- 20.73 nM and 251.2 +/- 55.25 nM, respectively. Site-directed mutagenesis of Trp-145, a residue postulated to contribute to the sugar-binding pocket in a manner akin to the essential Trp-183 in E. coli MglB, abolished Tp38's conformational change in response to D-glucose. The combined data are consistent with Tp38 serving as a glucose receptor for T. pallidum. These findings potentially have important implications for syphilis pathogenesis, particularly as they may pertain to glucose-mediated chemotactic responses by T. pallidum.