LT2, respectively, and was visualized by the Artemis Comparison Tool [57]. The gray areas indicate homologous regions with a minimum identity cutoff score of 88%. The region encoding acrD C646 cost is highlighted in light gray. The alignment was performed using the nucleotide search BLASTN from NCBI. (TIFF 1 MB) Additional file 4: Membrane protein topology of AcrD from Escherichia coli K-12 (A) and Erwinia amylovora Ea1189 (B). Description: The upper line indicates the predicted topology from TOPCONS [29] based on amino

acid sequences. Red lines indicate an inner membrane orientation; blue lines indicate an outer membrane orientation. Grey boxes indicate transmembrane helices spanning from the inside to the outside, white boxes indicate transmembrane helices spanning from the outside to the inside. Below the line is a graphical interpretation of the reliability of the prediction

for www.selleckchem.com/products/azd4547.html each amino acid. (TIFF 556 KB) Additional file 5: Scatter plot of the promoter activity of acrD from E. amylovora Ea1189. Description: It shows the effect of substrates on the promoter activity of acrD as determined by a transcriptional fusion with the reporter gene egfp. Antimicrobial compounds were added to cells of Ea1189 harboring pBBR.acrD-Pro.egfp by the 2-fold dilution method as described for MIC assays. EGFP fluorescence of the cells following exposure to various concentrations of the substrates was determined after 24 h incubation. A best-fit linear regression line between fluorescence and optical density values (dashed line) as well as a 95% confidence interval (solid line) are indicated. Outliers (black spots), showing higher fluorescence than the confidence interval, were identified as follows: deoxycholate (Doc), naringenin (Ng), tetracycline (Tc), and zinc sulfate Urocanase (Zn). The following substrates were applied to this assay: (+)-catechin, acridine orange, acriflavine, amikacin, azithromycin, benzalkonium chloride, berberine, bile salts, cadmium acetate, chloramphenicol, ciprofloxacin, clarithromycin, clotrimazol, cobalt chloride, copper sulfate, crystal violet, deoxycholate, erythromycin,

ethidium bromide, fusaric acid, fusidic acid, genistein, gentamycin, josamycin, luteolin, myricetin, naladixic acid, naringenin, nickel chloride, nitrofurantoin, norfloxacin, novobiocin, phloretin, polymyxin B, quercitin, rhodamine 6G, rifampicin, roxithromycin, SDS, silver nitrate, sodium arsenate, sodium tungstate, streptomycin, tetracycline, tetraphenylphosphonium chloride, tobramycin, and zinc sulfate. (TIFF 4 MB) Additional file 6: Primers used in this study. (DOCX 17 KB) References 1. Vanneste J: Fire blight: The disease and its causative agent, Erwinia amylovora. Oxon, UK: CABI Publishing; 2000.CrossRef 2. Bubán T, Orosz-Kovács ZS, Farkas Á: The nectary as the primary site of infection by Erwinia amylovora (Burr.). Plant Syst Evol 2003, 238:183–194. 3.