GLPG0259 free base is poorly soluble in aqueous media, and its so

GLPG0259 free base is poorly soluble in aqueous media, and its solubility decreases with increasing pH (<0.01 mg/mL at pH 7). Two approaches were developed in parallel to overcome this low solubility and to improve compound bioavailability after dosing in a solid dosage form. The first approach was a salt screening, AZD5363 which resulted in the selection of the fumarate salt for further formulation development work. The water solubility of

the GLPG0259 fumarate salt, as compared with that of the free base, was increased to 1.9–2.7 mg/mL. The impact of the improvement in solubility was confirmed in a comparative bioavailability study in fasted dogs. In that study, GLPG0259 fumarate salt (suspension in 20% [w/v] hydroxypropyl-ß–cyclodextrin, pH 3, or as crystalline powder in capsule form) resulted in plasma exposure similar to that of GLPG0259 free base in suspension in 20% acidified hydroxypropyl-ß–cyclodextrin, but 4-fold higher plasma exposure than that of GLPG0259 free-base crystalline powder in capsule form (data not shown). In humans, administration of GLPG0259 fumarate salt as a crystalline powder in capsule form leads to 50% lower bioavailability than that of GLPG0259 free base

given as a solution in 40% (w/v) hydroxypropyl-ß–cyclodextrin, pH 3 (study 3). The lower performance of the fumarate capsule in humans than in dogs is explained by the higher percentage of hydroxypropyl-ß–cyclodextrin (40% versus 20%) in the liquid formulation, which enhances GLPG0259 free-base solubility. Bafilomycin A1 mouse Concomitant food intake with the solid

dosage form Sitaxentan prevents this decrease in bioavailability by increasing the solubility further. The second approach was the improvement of GLPG0259 solubility by physical modifications of the drug substance – in particular, the development of solid dispersion formulations with GLPG0259 free base in an amorphous form homogenously dispersed in a polymer matrix. The free-base solid dispersion as a powder or pellets filled into capsules was tested in fasted dogs, and both solid dispersion formulations showed GLPG0259 plasma exposure similar to that of the fumarate salt as a crystalline powder in capsule form. Similar results were obtained in humans (study 4). In the Biopharmaceutical Classification System, drugs are classified according to measurements of solubility and permeability.[20] Regarding GLPG0259, it is a poorly soluble compound, with solubility that decreases with increased pH. The absorption of GLPG0259 was not measured in vivo in humans (there are no data after intravenous dosing), but its permeability assessed using the well established in vitro system, based on the human adenocarcinoma cell line Caco-2, was good, with an apparent permeability coefficient (Papp) of 12.4 10-6 cm/s and limited efflux (Papp B2A/Papp A2B = 2).

Curr Opin Infect Dis 16:129–134PubMedCrossRef Murphy TF, Brauer A

Curr Opin Infect Dis 16:129–134PubMedCrossRef Murphy TF, Brauer AL, Sethi S, Kilian M, Cai X, U0126 supplier Lesse AJ (2007) Haemophilus haemolyticus: a human respiratory tract commensal to be distinguished from Haemophilus influenzae. J Infect Dis 195:81–89PubMedCrossRef Musk DJ, Hergenrother PJ (2006) Chemical countermeasures for the control of bacterial biofilms: effective compounds and promising targets. Curr Med Chem 13:2163–2177PubMedCrossRef National Committee for Clinical Laboratory Standards (2000) Approved standard: M2-A7. Performance standards for antimicrobial disk susceptibility tests, 7th ed. National Committee

for Clinical Laboratory Standards, Wayne, Pennsylvania 19087-1898, USA National Committee

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54 Sulakvelidze A, Morris JG: Bacteriophages as therapeutic agen

54. Sulakvelidze A, Morris JG: Bacteriophages as therapeutic agents. Ann Med 2001, 33:507–509.PubMedCrossRef 55. Ritz HL, Kirkland JJ, Bond GG, Warner EK, Petty GP: Association of high levels

of serum antibody to staphylococcal toxic shock antigen with nasal Z-VAD-FMK nmr carriage of toxic shock antigen producing strains of Staphylococcus aureus . Infect Immun 1984, 43:954–958. 56. Kaliner MA: Human nasal respiratory secretions and host defense. Am Rev Respir Dis 1991, 144:S52–S56.PubMed 57. Rigby KM, DeLeo FR: Neutrophils in innate host defense against Staphylococcus aureus infections. Semin Immunopath 2012, 34(2):237–259. Competing interests The authors declare that they have no competing interests. Authors’ contributions SC, SK: Conceived and designed the experiments; PG: Performed the experiments; SC, SK: Analyzed the data; SC, SK: Wrote the paper. All authors read and approved the final manuscript.”
“Background The essential trace elemental selenium (Se) is the 34th element on the periodic selleck kinase inhibitor table and plays a fundamental role in human health [1]. Se is involved in several major metabolic pathways,

such as thyroid hormone metabolism, antioxidant defense systems and immune function [2]. In humans, selenium has navigated a narrow range from dietary deficiency (<40 μg per day) to toxic levels (>400 μg per day) [3]. Selenium toxicity in humans has been reported in the Chinese provinces Hubei and Shaanxi and in Indian Punjab, where Se levels in locally produced foods were found to be very high (750–4990 μg per person and day) [4]. The variation of Se status in humans both related to either Se excess or deficiency largely depends on the diet consisting of various crops, GNAT2 vegetables, fruits and meat [1]. Therefore, it is essential to understand the factors controlling the dynamic distribution of Se in the environment. Microorganisms

are involved in the transformation of selenium from one oxidation state to another [5-7]. A few studies reported that bacteria oxidized selenium to Se(IV) and Se(VI) in soils [8,9]. The formation of volatile methylated selenium species was also studied in several bacteria [5,7,10]. In addition, numerous bacteria were shown to reduce Se(VI)/Se(IV) to elemental Se, visible as red-colored nano-selenium [11-16]. Se(IV)-reducing bacteria generate red-colored elemental selenium nanoparticles (SeNPs) either under aerobic or under anaerobic conditions. Anaerobic Se(IV)-reducing bacteria encompass Thauera selenatis [17], Aeromonas salmonicida [18] and purple non-sulfur bacteria [14]. Aerobic bacteria involved in Se(IV) reduction include diverse species such as Rhizobium sp. B1 [19], Stenotrophomonas maltophilia SeITE02 [11], Pseudomonas sp. CA5 [13], Duganella sp. and Agrobacterium sp. [20]. However, the exact mechanism of selenium metabolism and reduction is still far from being elucidated.

Appl Phys Lett 1998,73(7):918–920 CrossRef 10 Jo SH, Huang ZP, T

Appl Phys Lett 1998,73(7):918–920.CrossRef 10. Jo SH, Huang ZP, Tu Y, Carnahan DL, Wang DZ: Effect of length and spacing of vertically aligned carbon nanotubes on field emission properties. Appl Phys Lett 2003,82(20):3520–3522.CrossRef 11. Jha A, Banerjee D, Chattopadhyay KK: Improved field emission from amorphous carbon

nanotubes by surface functionalization with stearic acid. Carbon 2011,49(4):1272–1278.CrossRef ABT263 12. Hazra KS, Gigras T, Misra DS: Tailoring the electrostatic screening effect during field emission from hollow multiwalled carbon nanotube pillars. Appl Phys Lett 2011,98(12):123116.CrossRef 13. Zhang YA, Wu CX, Lin JY, Lin ZX, Guo TL: An improved planar-gate triode with CNTs field emitters by CHIR-99021 clinical trial electrophoretic deposition. Appl Surf Sci 2011,257(8):3259–3264.CrossRef 14. Sanborn G, Turano S, Collins P, Ready WJ: A thin film triode type carbon nanotube field emission cathode. Appl Phys A 2013,110(1):99–104.CrossRef 15. Chen G, Neupane S, Li W, Chen L, Zhang J: An increase in the field emission from vertically aligned multiwalled carbon nanotubes caused by NH 3 plasma treatment. Carbon 2013, 52:468–475.CrossRef 16. Futaba DN, Kimura H, Zhao B, Yamada T, Kurachi H, Uemura S, Hata K: Carbon nanotube loop arrays for low-operational

power, high uniformity field emission with long-term stability. Carbon 2012,50(8):2796–2803.CrossRef 17. Pandey A, Prasad A, Moscatello JP, Engelhard M, Wang C, Yap YK: Very stable electron field emission from strontium titanate coated carbon nanotube matrices with low emission thresholds. ACS Nano 2013,7(1):117–125.CrossRef 18. Bonard BJ, Weiss N, Kind H, Stöckli T, Forró L, Kern

K: Tuning the field emission properties of patterned carbon nanotube films. Adv Mater 2001,13(3):184–188.CrossRef 19. Dolbec R, Irissou E, Chaker M, Guay D, Rosei F, El Khakani MA: Growth dynamics of pulsed laser deposited Pt nanoparticles on highly oriented pyrolitic graphite substrates. Phys Rev B 2004,70(20):201406.CrossRef 20. Aïssa B, Therriault D, El Khakani MA: On-substrate growth of single-walled carbon nanotube networks by an “all-laser” processing route. Carbon 2011,49(8):2795–2808.CrossRef 21. Collazo R, Schlesser R, Sitar Z: Role of adsorbates in field emission from nanotubes. Diam Relat Mater 2002, 211:769–773.CrossRef 22. Bower C, Zhu W, Jin Idoxuridine S, Zhou O: Plasma-induced alignment of carbon nanotubes. Appl Phys Lett 2000,77(6):830–832.CrossRef 23. Fowler RH, Nordheim L: Electron emission in intense electric fields. Proc Roy Soc A Math Phys Char 1926,119(781):173–181.CrossRef 24. Ago H, Kugler T, Cacialli F, Salaneck WR, Shaffer MSP, Windle AH, Friend RH: Work functions and surface functional groups of multiwall carbon nanotubes. J Phys Chem B 1999,103(38):8116–8121.CrossRef 25. Su W, Leung T, Chan C: Work function of single-walled and multiwalled carbon nanotubes: first-principles study. Phys Rev B 2007,76(23):235413.CrossRef 26.

Values are presented as means ± standard deviation (n = 36) * vs

Values are presented as means ± standard deviation (n = 36). * vs. rest, P < 0.001; # vs. After-exercise, P < 0.01. Glycogen concentrations in the tissues The glycogen concentration in the liver did not differ between the groups at any of the time points Everolimus cost (Figure 4A). Furthermore, the glycogen concentration in the white gastrocnemius muscle tissue did not differ between the groups at the rest and immediately post-exercise time points; however, this variable was significantly higher in the SP group than in the CON group at the recovery period time point (1 h post-exercise; Figure 4B). In contrast, no

significant between-group differences were observed in the red gastrocnemius muscle tissue (Figure 4C). Figure 4 Changes in the glycogen levels during exercise and after 1 h of exercise. CON: distilled water LGK-974 with training, SP: silk peptide-treated with training. A, liver; B, white gastrocnemius muscle tissue; and C, red gastrocnemius muscle tissue at rest, after exercise, and recovery in the CON and SP groups. Values are presented as means ± standard deviations (n = 36). * vs. rest, P < 0.01; # vs. rest and after-exercise, P < 0.05; $ vs. recovery in CON, P < 0.001; ¶ vs.

after-exercise, P < 0.05. Discussion The present study demonstrated that a 2-week regimen of silk peptide (SP) treatment and endurance training could increase the max, whereas endurance training alone had no similar effect. Rebamipide A 2-week period of SP treatment also increased fat oxidation during the initial phase of exercise in exercised mice. In human studies, the max test during

graded treadmill exercise is the most commonly used endurance performance measurement [20, 21]. In the present study, max was not changed in the CON group after the 2-week training. Our previous study demonstrated that max was significantly increased by 4 week-training which the intensity was the same with the present study training protocol [16]. Thus, the duration (2 weeks) and/or intensity (75% of VO2 max) seem not to be enough to increase the endurance capacity in the present study. On the other hand, the max was significantly increased after a 2-week period of SP treatment when compared with the same metric before training. A previous study reported that a 30-day SP treatment regimen (800 mg/kg body weight daily) and swimming exercise training increased the maximum swimming time of mice by reducing exercise-induced tissue injuries and energy depletion [13]. In addition, a 44-day SP treatment regimen led to an increased maximum swimming time and decrease in the levels of muscle tissue damage markers such as creatine kinase, aspartate aminotransferase, and lactate dehydrogenase in a dose-dependent (50, 160, and 500 mg/kg) manner after forced swimming exercises [12]. Therefore, it seems that SP treatment can increase the exercise capacity regardless of the type of exercise.

e , misclassification does not depend on cohort), the study resul

e., misclassification does not depend on cohort), the study results for the measure of nonvertebral sites and for vertebral sites are likely more attenuated by misclassification than results at the hip. In conclusion, for this large

observational study of more than 200,000 bisphosphonate patients, the apparent differences in the baseline incidence of hip fractures among the alendronate, risedronate, and ibandronate cohorts likely reflect differences in the risk profile of patients prescribed each bisphosphonate. Statistical adjustments could not account for these differences and therefore the design of epidemiological studies should be SCH772984 mouse given careful consideration to account for these differences. Relative to the baseline fracture incidence, the longitudinal analyses indicated that alendronate and risedronate decreased nonvertebral and hip fractures over time, whereas ibandronate did not. All three bisphosphonates decreased vertebral fractures. The reductions Tyrosine Kinase Inhibitor Library cell assay observed in fracture incidence over time within each cohort suggest that the effectiveness

of each bisphosphonate in clinical practice has been consistent with their efficacies demonstrated in randomized controlled trials. Acknowledgement Funding by The Alliance for Better Bone Health (Procter & Gamble Pharmaceuticals and sanofi-aventis). Conflicts of interest Dr. Abelson reports receiving consulting fees from sanofi-aventis, Procter & Gamble, Novartis; serving on speaker’s bureaus for Amgen, Procter & Gamble, Roche, Novartis, and sanofi-aventis. Dr. Gold reports receiving consulting or advisory committee fees from Amgen, Eli Lilly, GlaxoSmithKline, Merck, Procter & Gamble, Roche, sanofi-aventis; serving on

Glycogen branching enzyme speaker’s bureaus for Amgen, Eli Lilly, GlaxoSmithKline, Procter & Gamble, Roche, and sanofi-aventis. Dr. Thomas reports receiving consulting or advisory committee fees from Amgen, Daïchi-Sankyo, Ipsen, Lilly, MSD, Novartis, Procter & Gamble, Roche/GlaxoSmithKline, sanofi-aventis, and Servier; grant support from Lilly, MSD, Nicomed, Novartis, Procter & Gamble, sanofi-aventis, and Servier. Dr. Lange is an employee of Procter & Gamble. Open Access This article is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited. References 1. Avorn J (2007) In defense of pharmacoepidemiology—embracing the yin and yang of drug research. N Engl J Med 357:2219–2221CrossRefPubMed 2. Perreault S, Dragomir A, Blais L et al (2008) Population-based study of the effectiveness of bone-specific drugs in reducing the risk of osteoporotic fracture. Pharmacoepidemiol Drug Saf 17:248–259CrossRefPubMed 3. Langsetmo LA, Morin S, Richards JB et al (2009) Effectiveness of antiresorptives for the prevention of nonvertebral low-trauma fractures in a population-based cohort of women. Osteoporos Int 20:283–290CrossRefPubMed 4.

Analysis of microarray images was carried out applying the ImaGen

Analysis of microarray images was carried out applying the ImaGene 6.0 software (BioDiscovery) as described previously [42]. Lowess normalization and significance test (fdr) were performed with the EMMA software [60]. M-values (log2 experiment/control ratio), P-values (t test) and A-values were also calculated with EMMA. The M-value represents the logarithmic ratio between both channels. The A-value represents the logarithm of the combined intensities of both channels. The microarray Caspase inhibitor results were verified for specific genes

by quantitative reverse transcription-PCR using a QuantiTect SYBR Green reverse transcription-PCR kit (QIAGEN, Hildesheim, Germany) according to the manufacturer’s instructions. Filtering find more and clustering analysis of the microarray data K-means

clustering analysis of the microarray time-course data was performed with the aid of the Genesis software [62]. After normalization, only genes with approximately threefold change in expression (M-value of ≥ 1.4 or ≤ -1.4) in at least one point of time in the wild type microarrays were considered for clustering analysis. Genes that did not present an evaluable expression value for at least 5 of the 6 points of time (missing values on the microarray flagged as empty spots) were not considered. K-means clustering was used for distributing differentially regulated genes into 6 groups, both with the wild type and with the rpoH1 mutant microarray data. Quantitative RT-PCR analyses Reverse transcription

was performed using Superscript II reverse transcriptase (Invitrogen) with random hexamers as primers. RNA samples were tested for two time points, 10 and 60 minutes after pH shock. Real-time PCRs were run on an Opticon system (BioRad) using the FastStart DNA MasterPLUS SYBRGreen I kit (Roche) according to the manufacturer’s instructions. The housekeeping gene rkpK was used as a reference for normalization. The sequences of the primers used are available at http://​www.​cebitec.​uni-bielefeld.​de/​groups/​brf/​software/​gendb_​info/​. Three independent cultures were analyzed, as GPX6 well as three technical replicates, for each time point. Microarray data accession numbers The entire set of microarray data has been deposited in the ArrayLims database [63]. Acknowledgements We thank Victoria Gödde, Manuela Meyer and Eva Schulte-Berndt for providing outstanding technical help. This work was supported by a scholarship from the NRW Graduate School in Bioinformatics and Genome Research, funded by the Ministry of Innovation, Science, Research and Technology of the state of North Rhine-Westphalia, Germany. Electronic supplementary material Additional file 1: Complementation of rpoH1 mutation.

Blunt JW, Copp BR, Hu W-P, Munro MHG, Northcote PT, Prinsep MR: M

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starter unit. Gene 2002, 296:235–247.CrossRefPubMed 5. Chang Z, Sitachitta N, Rossi JV, Roberts MA, Flatt PM, Jia J,

Sherman DH, Gerwick WH: Biosynthetic pathway and gene cluster analysis of curacin A, an antitubulin natural product from the tropical marine cyanobacterium Lyngbya majuscula. J Nat Prod 2004, 67:1356–1367.CrossRefPubMed 6. Edwards DJ, Marquez BL, Nogle LM, McPhail K, Goeger DE, Roberts MA, Gerwick WH: Structure and biosynthesis of the jamaicamides, Protein Tyrosine Kinase inhibitor new mixed polyketide-peptide neurotoxins from the marine cyanobacterium Lyngbya majuscula. Chem Biol 2004, 11:817–833.CrossRefPubMed 7. Gu L, Geders TW, Wang B, Gerwick WH, Håkansson K, Smith JL, Sherman DH: GNAT-like strategy for polyketide chain initiation. Science 2007, 318:970–974.CrossRefPubMed 8. Cragg GM,

Newman DJ, Snader KM: Natural products in drug discovery and development: The United States National Cancer Institute Role. Phytochemicals in Human Health Protection, Nutrition, and Plant Defense (Edited by: Romeo JT). New York: Kluwer Academic/Plenum Publishers 1999, 1–29. 9. Suyama TL, Gerwick WH: Stereospecific total synthesis of somocystinamide A. Org Lett 2008, 10:4449–4452.CrossRefPubMed 10. Pfeifer BA, Wang CCC, Walsh CT, Khosla C: Biosynthesis of yersiniabactin, a complex polyketide-nonribosomal peptide, using Escherichia coli as a heterologous host. Appl Environ Microbiol 2003, 69:6698–6702.CrossRefPubMed 11. Schmidt EW, Nelson JT, Rasko DA, Sudek S, Eisen JA, Haygood MG, Ravel J: Patellamide A and C biosynthesis by a microcin-like pathway in Prochloron didemni , the cyanobacterial symbiont of Lissoclinum patella. Proc Natl Acad Sci enough USA 2005, 102:7315–7320.CrossRefPubMed 12. Watanabe K, Hotta K, Praseuth AP, Koketsu K, Migita A, Boddy CN, Wang CCC, Oguri H, Oikawa H: Total biosynthesis of antitumor nonribosomal peptides in Escherichia coli. Nat Chem Biol 2006, 2:423–428.CrossRefPubMed 13. Wilkinson B, Micklefield J: Mining and engineering natural-product biosynthetic pathways. Nat Chem Biol 2007, 3:379–386.CrossRefPubMed 14. Galm U, Shen B: Expression of biosynthetic gene clusters in heterologous hosts for natural product production and combinatorial biosynthesis. Exp Op Drug Discov 2006, 1:409–437.CrossRef 15.

Cell proliferation occurred after

Cell proliferation occurred after BGJ398 molecular weight 2~3 days of culture in the ATRA/growth factor group. The cell growth in this group was almost the same as in the growth

factor group, but the number and volume of the cell spheres formed were slightly smaller than those in the growth factor group. Cell proliferation also occurred after 2~3 days in the ATRA group, with the cell spheres exhibiting suspended growth, but only cell masses consisting of dozens of cells were observed during the whole process. The volume of the cell spheres was larger than that in the control group, but obviously smaller than that in the growth factor group and the ATRA/growth factor group. The cell proliferation in the control group was relatively slower, and the formed colonies were smaller, merely consisting of a dozen cells (Fig. 3). No obvious adherent differentiation was observed in any group. With the mean of optical density values measured for each group as the vertical axis, and the growth days as the horizontal axis, the growth curves of BTSCs for different groups were plotted (Fig. 4) to

compare the cell proliferation rates of the four groups. It can be observed that, on the 1st-3rd day, the growth curves of all the four groups rise slowly, with an insignificant difference in the cell proliferation rate. From the 3rd day, the cell proliferation obviously become Small molecule library molecular weight more rapid, and the growth curves of the four groups begin to separate from each other. The curve is steep during the 5th~7th days, indicating the peak of proliferation. Cell proliferation is slowest in the control group, obviously faster in the ATRA group, and fastest in the growth factor group, and the proliferation rate of the ATRA/growth factor group is slightly lower than that of the growth factor group, but significantly higher than that of the ATRA group. It is indicated that ATRA had a promotive effect on the proliferation of suspended BTSCs, but had no obvious synergistic or antagonistic effect with

the growth factor. Figure 3 The volume of the cell spheres Methocarbamol formed in different group(Inverted phase-contrast microscope, × 400). 2A: the control group. 2B: the ATRA group. 2C: the ATRA/growth factor group. 2D: the growth factor group. Figure 4 Growth curves of BTSCs in different groups(the mean of optical density values measured for each group as the vertical axis, and the growth days as the horizontal axis). The results are shown as mean ± SD of four different experiment. Data of each day was analyzed by one-way ANOVA with Dunnett t test. The growth curves of the ATRA group, ATRA/growth factor group and growth factor group rise faster than that of the control group(P < 0.01). While there were no statistically significant between the ATRA/growth factor group and growth factor group(P > 0.05).

These

These DAPT nmr amino acids were changed into either a phenylalanine (F) residue that cannot become phosphorylated or an aspartate (D) residue to mimic a modification resulting in an additional negative charge. All constructs were functionally active, i.e. AI-2 was still produced by these modified proteins (data not shown). Total protein lysates of S. Typhimurium luxS mutant strains containing one of these point mutated LuxS constructs, were analyzed with 2D gel electrophoresis (2DE). As shown in Figure 2D-F, all strains with Y to

F mutations still possess two LuxS spots. This rules out any of the tyrosine residues as target sites for modification. Furthermore, the pI shift seen in the Y to D mutation strains (Figure 2G-I) confirms the charge difference on the modified LuxS form. This result also illustrates that the interpretation of proteomic results has to be done with great care. Posttranslational modifications all correspond to a specific shift in pI and/or molecular weight. In this respect, we suggest that the postulated phosphorylation of LuxS in Bifidobacterium longum proposed by Yuan et al. should be re-investigated [22]. Figure 2 2DE analysis of Salmonella Typhimurium luxS mutants. (A) Total gel image of wildtype S. Typhimurium proteins. The two LuxS forms are indicated with an arrow. Based

on pI calculations, the right spot corresponds to native LuxS and the left spot carries a posttranslational modification. this website (B-J) Close-up view of the area of the LuxS spots in a luxS mutant carrying different LuxS complementation constructs. (B) negative control – empty vector; (C) wildtype LuxS; (D) LuxS-Y88F; (E) LuxS-Y126F; (F) LuxS-Y131F; (G) LuxS-Y88D; (H) LuxS-Y126D; (I) LuxS-Y131D; (J) LuxS-C83A. Remark that in theory, on the gels from which panels Phospholipase D1 G-I are taken, an additional modified LuxS spot is expected, accumulating the Y to D mutation and the cysteine modification.

For Bacillus subtilis LuxS, oxidation of C84 has previously been reported with purified LuxS protein in studies to reveal the reaction mechanism of the synthase [23–25]. This oxidation is irreversible and adds one negative charge to the protein [23], which makes it a good candidate for the LuxS modification we detected in the S. Typhimurium proteome. Analogous to the tyrosine mutant constructs, we made a point mutation of the corresponding cysteine residue in S. Typhimurium to an alanine residue (C83A) which can no longer be oxidized and subsequently analyzed this strain by 2DE. As shown in Figure 2J the C83A luxS strain lacks the acid shifted LuxS spot confirming C83 as the target for posttranslational modification. As this cysteine residue is required for LuxS catalytic activity [26], the LuxSC83A mutant strain failed to produce AI-2 as revealed by the use of the AI-2 bioassay [27] (data not shown).