Tauro β muricholic acid

Taurocholic acid (TCA), tauro-β-muricholic acid (TMCA), tauromurideoxycholic acid (TMDCA), taurochenodeoxycholic acid (TCDCA), taurodeoxycholic acid (TDCA), taurolithocholic acid (TLCA), cholic acid (CA), α-muricholic acid (αMCA), β-muricholic acid (βMCA), ω-muricholic acid (ωMCA), murideoxycholic acid (MDCA), ursodeoxycholic acid (UDCA), chenodeoxycholic acid (CDCA), deoxycholic acid (DCA), and lithocholic acid (LCA) were purchased from either Steraloids, Inc. (Newport, RI) or Sigma-Aldrich (St Louis, MO). Clofibric acid (CLOF) was purchased from Sigma-Aldrich Co. (St. Louis, MO). Chloroform, agarose, and ethidium bromide were purchased from AMRESCO Inc. (Solon, OH). RNA-Bee was purchased from TelTest Inc. (Friendswood, TX). All other chemicals, unless indicated, were purchased from Sigma-Aldrich Co. (St. Louis, MO).

P_tcpA-sh-ble was grown as overnight culture, diluted 1:1000 in fresh LB, and grown for ~2 hours at 37°C. Each reaction was prepared in 40μl 0.5X pH 8.5 LB medium, with sodium taurocholate hydrate (TC, Sigma-Aldrich), sodium glycocholate hydrate (Sigma Aldrich), cholic acid (CA, Alfa Aesar), sodium taurodeoxycholate hydrate (Sigma-Aldrich), sodium glycodeoxycholate (Sigma Aldrich), deoxycholic acid (DCA, MB Biomedicals), tauro-β-muricholic acid (Steraloids Inc.), or β-muricholic acid (Steraloids Inc.) added to a final concentration of 125μM. 2μl of reporter strain subculture was then added, and samples incubated anaerobically at 37°C for 4hrs. 2μl of each reaction was then added to 200μl of 0.5X LB pH 8.5 +/−10μg/ml of zeocin (Sigma Aldrich), incubated for 30 minutes aerobically at 37°C with agitation, and then serially diluted and plated onto LB agar plates with streptomycin to determine survival rates. Induction represents percentage of P_tcpA-sh-ble reporter cells surviving treatment with zeocin after incubation with indicated samples under anaerobic conditions, defined as (zeocin-treated sample survival/average of no-zeocin controls)*100. Where noted, survival rates were normalized to that induced by 125μM taurocholate.