Similarly one can show that the F(t)/F o response changes (blue solid curve) when the rate constant of the release of DSQ is assumed to be 50-fold higher with k dsq~ 15 μs−1, which would mean the ignorance of DSQ release in a time domain above ~10 μs. Fig. 1 Relative chlorophyll a fluorescence change (closed black diamonds) F(t)/F o of 1 h dark-adapted Arabidopsis thaliana leaf in 100 ns to 10 s time range (logarithmic) upon saturating laser flash (6.2 × 1015 photons cm−2/flash), reproduced from Fig. 2 in Steffen et al.

(2005). Bold red curve is the simulated response F DSQ(t) using a modification of Eq. 1a. The modification accounts for a S 0 (β):S 1:S 2 heterogeneity of 0.2:0.4:0.4 with corresponding rate constants of donor side quenching k dsq = 300, 60, and 7 ms−1, k AB~9 ms−1 and a biphasic decay of QB-nonreducing RCs with rate constants k −nqb~25 and 0.5 s−1 Selleckchem MK5108 and nF v = 1.8. Note that F pl is from (reduced) QB-nonreducing RCs at the fractional size β ~ 0.3/1.8~18%. The red dashed curves (closed triangles, diamonds and squares) are simulations with variable rate constant of quenching recovery (k AB) due to Q A − reoxidation. Parameter values of variable quenching-regeneration (k AB) are indicated at the right-hand side of the respective curves. The blue-colored check details dashed curve shows the F DSQ(t) response

when, at constant k AB (~10 ms−1), k dsq is increased 50-fold (for instance when donor side quenching (DSQ) is ignored). The dashed curves illustrate

the effect of interference between k dsq and k AB on the maximum of F(t)/F o with an increasing disproportion between n\( F_\textv^\textSTF (-)-p-Bromotetramisole Oxalate \) and the maximum of F DSQ(t) with the increase in rate (k AB) of quenching recovery In summary, the quantitative data on laser flash-induced variable fluorescence from the 100 ns to 1 ms time range (Belyaeva et al. 2008) confirming those of others (Steffen et al. 2001, 2005; Belyaeva et al. 2006), need a substantial correction with respect to magnitude of the normalized variable fluorescence associated with single turnover-induced charge separation in RCs of PS II. Their data are conclusive with the involvement of donor side quenching, the release of which occurs with a rate constant in the range of tens of ms−1, and presumed to be associated with reduction of \( Y_\textz^ + \) by the OEC. Open Access This article is distributed under the terms of the Creative Commons Attribution click here noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited. References Belyaeva NE, Paschenko VZ, Renger G, Riznichenko GYu, Rubin AB (2006) Application of photosystem II model for analysis of fluorescence induction curves in the 100 ns to 10 s time domain after excitation with a saturating light pulse.

Infect Immun 2006, 74:2154–2160.PubMedCrossRef 32. Hodzic E, Borjesson DL, Feng S, Barthold SW: Acquisition dynamics of Borrelia burgdorferi and the agent of human granulocytic ehrlichiosis at the host-agent interface. Vector Borne Zoonotic Dis 2001, 1:149–158.PubMedCrossRef 33. Kung F, Anguita J, Pal U: Borrelia burgdorferi and tick proteins supporting pathogen persistence in the vector. Future Microbiol 2013, 8:41–56.PubMedCrossRef 34. Armstrong AL, Barthold SW,

Persing DH, Beck DS: Carditis in Lyme disease susceptible and resistant strains of laboratory mice infected with Borrelia burgdorferi . Am J Trop Med Hyg 1992,47(2):249–258.PubMed 35. Barthold SW, Sidman CL, Smith AL: Lyme borreliosis in genetically resistant and susceptible mice with severe combined immunodeficiency. Am J Trop Med Hyg 1992, 47:605–613.PubMed 36. Casjens PRN1371 S, Palmer N, van Vugt R, Huang WM, Stevenson B, Rosa P, Lathigra R, Sutton G, Peterson J, Dodson RJ, et al.: A bacterial genome in flux: the twelve linear and nine circular extrachromosomal DNAs in an infectious isolate of the Lyme disease spirochete Borrelia burgdorferi . Mol Microbiol 2000, 35:490–516.PubMedCrossRef 37. Fraser CM, Casjens S, Huang WM, Sutton GG, Clayton R, Lathigra R, White O, Ketchum KA, Dodson R, Hickey EK, et al.: Genomic sequence

of a Lyme disease spirochaete, Borrelia burgdorferi . Nature see more 1997, 390:580–586.PubMedCrossRef 38. Grimm D, Eggers CH, Caimano MJ, Tilly K, Stewart PE, Elias AF, Radolf JD, Rosa PA: Experimental assessment of the roles of linear plasmids lp25 and lp28–1

of Borrelia burgdorferi throughout the infectious cycle. Infect Immun 2004, 72:5938–5946.PubMedCrossRef Mannose-binding protein-associated serine protease 39. Barbour AG: Isolation and cultivation of Lyme disease spirochetes. Yale J Biol Med 1984, 57:521–525.PubMed 40. Samuels DS, Mach KE, Garon CF: Genetic buy BLZ945 transformation of the Lyme disease agent Borrelia burgdorferi with coumarin-resistant gyrB. J Bacteriol 1994, 176:6045–6049.PubMed 41. Ohnishi J, Piesman J, de Silva A: Antigenic and genetic heterogeneity of Borrelia burgdorferi populations transmitted by ticks. Proc Natl Acad Sci USA 2001, 98:670–675.PubMedCrossRef 42. Barthold SW, Persing DH, Armstrong AL, Peeples RA: Kinetics of Borrelia burgdorferi dissemination and evolution of disease following intradermal inoculation of mice. Am J Pathol 1991, 139:263–273.PubMed 43. Reed LJ, Muench H: A simple method of estimating fifty per cent endpoints. Am J Hyg 1938, 27:493–497. Competing interests The authors declared that they have no competing interests. Authors’ contributions DI, KH, EH and SWB performed and analyzed results. SF, EH and SWB participated in experimental design. DI, KH, EH and SWB co-wrote the manuscript. All authors read and approved the manuscript.

The semi-quantitative evaluation of both the absolute values of the apposition bands and the width of daily bone apposition values increased for treated compared to untreated rats, but these effects were not significant. Table 2 Results of the intravital fluorochrome labeling   SHAM SHAM Vib. OVX OVX Vib. OVX vs. SHAM Vib vs. non vib Mean STD Mean STD Mean STD Mean STD p value p value Absolute apposition bandwidth (m -6 ) Calcein green (d0 − d18) 696 275 822 226 1093 182

1032 290 <0.0001 0.4829 Alizarin red (d18–d24) 823 271 804 229 889 181 944 274 0.0267 0.6943 Tetracycline (d24–d35) 659 333 641 226 669 219 709 242 0.4267 0.8278 Sum 2,178 2,267 2,651 2,685             Absolute apposition bandwidth per day (m –6 ) Calcein green (d0–d18) 38.6 15.3 45.7 NVP-HSP990 price 12.5 60.7 10.1 57.3 16.1 <0.0001 0.4877 Alizarin Thiazovivin manufacturer Red (d18–d24) 137.2 45.2 134.1 38.2 148.2 30.2 157.3 45.6 0.0269 0.7024 Tetracycline (d24–d35) 59.9 30.3 58.3 20.5 60.8 19.9 64.5 22.0 0.4275 0.8227 Sum 235.7 238.1 269.7 279.1             Relative apposition bandwidth per day (%) Calcein green (d0–d18) 16.8 4.0 19.4 3.4 22.9 3.9 20.7 2.7 <0.0001 0.7371 Alizarin red (d18–d24) 58.5

5.0 56.3 4.7 54.9 3.3 56.2 6.1 0.0436 0.6052 Tetracycline (d24–d35) 24.7 7.0 24.3 4.8 22.2 4.0 23.2 5.5 0.0831 0.8085 The p value of the difference between treated and untreated animals was calculated using a two-way ANOVA. p values <0.05 were considered significant Flat-panel volumetric computed tomography The SHAM group had

a significantly ARRY-438162 order improved BMD, cancellous and cortical bone density compared to OVX animals (p < 0.0001 for all). Vibration led to an improvement of total BMD, cancellous and cortical BMD (Table 1). The cortical bone density after vibration was significantly improved (p = 0.0035), while the BMD (p = 0.0532) and cancellous bone density (p = 0.0634) showed improvement; however, the improvement failed BCKDHB to reach significant values. The main disadvantage of the fpVCT used in this study was the lower spatial resolution compared to the µCT. The former method does not allow a detailed description of the trabecular microstructure. Ashing The ash-BMD of SHAM rats was significantly improved compared to OVX rats (p < 0.0001). Vibration yielded a significant improvement of ash-BMD in all groups (p = 0.0011). There were no differences between groups before ashing. After ashing, the SHAM-operated animals had higher ash weights compared to OVX, but these changes were not significant. After calculating the ash-BMD, more differences between the groups were observed (Table 1). Discussion Osteoporosis primarily affects trabecular bone. In humans, the majority of osteoporotic fractures occur in the spine and metaphysis of long bones. In the rat osteopenia model, osteoporosis mainly affects the metaphyseal tibia and lumbar spine [19–23].

, Uxbridge, UK) with 5.0 kV voltage and 10.0 μA current, on top and side views. After each heating stage, the specimens were scanned by home-made XPS. Core level and valance band photoelectron spectra were excited by monochromatic Al K radiation (1,487 eV) and collected, at take-off angle of 35°,

by a hemispherical analyzer with adjustable overall resolution between 0.8 and 1.2 eV. The surveys were conducted in various ranges of electron energies including the overall binding energy survey (0 to 1,000 eV) besides individual spectra for Si 2p (95.0 to GW786034 clinical trial 110.0 eV), C 1 s (282.0 to 287.0 eV) and O 1 s (520 to 550 eV) which were monitored more accurately in a discrete number of scans. All spectra were taken at room temperature in a UHV chamber of about 10−10 Torr pressure. The resulting XPS spectra were analyzed by spectral decomposition using the XPS peak software and their oxide levels were determined. Results and discussion The VLS-grown Si NWs used in this study Lazertinib cell line were randomly oriented with average diameter and length of 84.96 nm and 3.508 μm, respectively. The pristine Si NWs are covered by a native oxide layer of 1 to 4 nm. SEM and transmission electron microscopy (TEM) micrographs of the pristine Si NWs are depicted in Figure 1. Residual gold nanoparticles

were removed by rinsing the Si NWs into HNO3 solution preventing its catalytic effect on oxidation. Figure 1 SEM and transmission electron microscopy (TEM) micrographs of the pristine Si NWs. (a) Top-view SEM micrograph of the Si NWs grown by VLS mechanism showing their random orientation. (b) TEM image of an individual Si NW cross-section representing the continuous native oxide layer of 3 to 4 nm in diameter atop. Regarding the micrographs, the Si core diameter can be estimated as 50 ± 10 nm. The red dotted line

insists on the fact that TEM micrograph is taken for a NCT-501 mw single Si NW among the large ensemble observed through SEM. As an illustrative Si 2p spectrum of oxidized Si NWs, the Si 2p spectrum of the H-terminated Si NWs annealed at 500°C for 60 min is depicted in Figure 2. By formation of even very thin silicon dioxide layers, the Si 2p XPS survey of Si NWs changes, showing a peak between the binding energies of 102 to 104 eV. To quantitatively evaluate PD184352 (CI-1040) the oxidation process, Si 2p spectral decomposition was conducted on the spectra after Shirley background subtraction, through a curve-fitting procedure using Gaussian-Lorentzian functions [16]. Consequently, the Si 2p spectra can be divided into six different sub-peaks including two silicon spin-splitting peaks as Si 2p 1/2 and Si 2p 3/2, three silicon sub-stoichiometric oxides (known as suboxides) peaks as Si2O, SiO and Si2O3, and the silicon dioxide (SiO2) peak. The chemical shifts (Δ) of the sub-peaks obtained in Figure 2 relative to the Si 2p 3/2 (at 99.60 ± 0.02 eV) are as follows: Si 2p 1/2 (Δ = 0.60 eV), Si2O (Δ = 0.97 eV), SiO (Δ = 1.77 eV), Si2O3 (Δ = 2.50 eV), and SiO2 (Δ = 3.87 eV).

e., the beam is directed through the fused silica substrate onto the SiO x film (Figure 1b). To determine the intensity distribution in the image plane on the sample, the sample is removed, and this plane is imaged onto a UV-sensitive CCD camera using a × 100 UV microscope objective (Ultrafluar, Carl Zeiss, Oberkochen, Germany) (Figure 1c). Irradiation experiments with high spatial resolution were carried out using a standard ArF excimer

laser emitting at 193 nm with pulse duration of about 20 ns. In this case, a Schwarzschild-type reflective objective (NA = 0.4, ×25 demagnification) was used for mask projection. A scanning electron microscope (Zeiss DSM 962) has been used to investigate XMU-MP-1 ic50 the laser-induced morphological changes. Results Figure 2 displays SiO x films irradiated with a crossed grating pattern with and without PDMS confinement layer (after peeling off selleck inhibitor this layer). In both cases, the film disintegrates with a buy AZD4547 period given by the beam pattern, whereas the fused silica substrate remains

intact. Confinement leads to smooth, contiguous features around the ablation sites instead of irregular splashes observed without this confinement. Figure 2 Influence of confinement. Patterned 150-nm-thick SiO x film irradiated (a) without and (b) with confinement (after peeling off the confinement layer); laser parameters: 248 nm, 260 mJ/cm2, 1 pulse. To establish a correlation between the irradiation pattern and the resulting grid pattern, beam profiles in the sample plane have been recorded (Figure 3). In the case of a large period of the mask (40 μm), the intensity pattern is a four times reduced, but congruent, image of the transmission pattern of the mask (a). In the case of the 20-μm mask period, the beam pattern is already Urocanase a bit blurred due to the limited resolution of the projection optics (f). The corresponding grid patterns obtained at various fluences are also displayed in Figure 3. At low fluence, in the case of a period large compared to the optical resolution, the film detaches from the substrate in the area of the irradiated cross

pattern forming hollow channels, but keeping contact to the substrate in the non-irradiated areas (b). For the smaller period, only some buckling of the film at the high intensity crossing points is observed (g). Increasing the fluence, after enlargement of the detached area (c, h), rupture of the film in between the crossing points of the channels and formation of openings in the detached film occur (d, i). At still higher fluence, the enlargement of the openings (e) and the formation of thin wires of residual material between these openings (k) are observed. However, at the positions of minimum intensity, this wire grid is still connected to the substrate. Depending on the fluence and the particular intensity pattern, other types of shaping can be observed, e.g., hollow channels or arrays of blisters or cup-like structures.

Microscopic agglutination test (MAT) The microscopic agglutination test was performed Luminespib nmr according to [1]. In brief, an array of 22 serovars of Leptospira spp. as antigens were employed: Australis, Autumnalis, Bataviae, Canicola, Castellonis, Celledoni, Copenhageni, Cynopteri, Djasiman, Grippotyphosa, Hardjo, Hebdomadis, Icterohaemorrhagiae, Javanica, Panama, Patoc, Pomona, Pyrogenes, Sejroe, Shermani, Tarassovi and Wolffi. All the strains were maintained in EMJH liquid medium (Difco, USA) this website at 29°C. A laboratory – confirmed case of leptospirosis was defined by the demonstration of a four – fold microagglutination titer rise

between paired serum samples. The probable predominant serovar was considered to be the one with the highest dilution that could cause 50% of agglutination. MAT was considered negative when the titer was below 100. Characterization of the protein in silico Predicted coding sequence (CDSs) LIC11834 and LIC12253 were identified on L. interrogans serovar Copenhageni and selection was based on cellular localization; cellular localization prediction was performed by PSORT, http://​psort.​nibb.​ac.​jp[54] and PredictProtein web server, https://​www.​predictprotein.​org/​[25]. The SMART [23]http://​smart.​embl-heidelbergde/​ and PFAM [55]http://​www.​sanger.​ac.​uk/​Software/​Pfam/​ web servers were used to search for predicted functional and structural domains. The presence

of lipobox putative sequence MK0683 datasheet was evaluated by use of the LipoP program [56]http://​www.​cbs.​dtu.​dk/​services/​LipoP/​. selleck screening library The predicted sequence of the lipobox was also assessed by use of the SpLip program, as described by Setubal

et al. [57]. Secondary structure, solvent accessibility and cellular localization predictions were also performed by using PredictProtein web server, https://​www.​predictprotein.​org/​[25]. DNA isolation and PCR analysis Leptospira cultures were harvested by centrifugation at 11,500 g for 30 min and gently washed in sterile PBS twice. Genomic DNA was isolated from the pellets by guanidine – detergent lysing method using DNAzol® Reagent (Invitrogen), according to the manufacturer’s instructions. Primers were designed according to L. interrogans serovar Copenhageni genome sequences (GenBank accession AE016823) and are listed in Table 1. PCR was performed in a reaction volume of 25 μl containing 100 ng of genomic DNA, 1 × PCR buffer (20 mM Tris – HCl, pH 8.4, 50 mM KCl), 2 mM MgCl2, 20 pmol of each specific primer, 200 μM of each dNTP, and 2.5 U Taq DNA Polymerase (Invitrogen). Cycling conditions were: 94 ° C – 4 min, followed by 40 cycles at 94°C – 50 sec, 57°C (LIC11834) or 56°C (LIC12253) – 50 sec, 72°C – 90 sec, and a final extension cycle of 7 min at 72°C. PCR amplified products were loaded on a 1% agarose gel for electrophoresis and visualization with ethidium bromide.

Because of the radius of neighboring crystal layers, the uncut thickness should be a range rather than a certain value, as displayed in Table 2. Figure 6 Displacement vector sum of each layer in y direction. Table 2 The uncut thickness in different combinations of depth of cut and lattice plane PFT�� price Cutting direction Cutting depth (nm) Uncut thickness (nm) on (010) surface 1 0.45-0.58 2 0.87-1.01 3 1.23-1.38 on (111) surface 1 0.35-0.58 2 0.68-0.93   3 1.07-1.28 Figure 7 shows the average uncut thickness in different undeformed chip thicknesses when machined surfaces are (010) and (111) plane,

respectively. The uncut thickness increases with an increase in undeformed chip thickness. With the same combination of cutting direction and crystal orientation, the uncut thickness is nearly proportional to the undeformed chip thickness Talazoparib clinical trial on our simulation scale [17]. The uncut thickness of machining on (010) crystal orientation is about 0.1 nm bigger than that on (111) crystal orientation with the same undeformed

chip thickness, which means that the difference can be ignored considering the interplanar distance. Figure 7 The uncut thickness. In different depths of cut when machined surfaces are (010) and (111) plane, respectively. Cutting force and energy The cutting force derives from the interaction between the tool and material atoms in the molecular dynamics simulation of nanometric cutting. Since it has a great influence on the surface finish, tool wear, etc., the cutting force is monitored during the machining process. The sum of force vector GDC-0449 purchase on three axes directions, namely Fx, Fy, and Fz, are defined as tangential force, normal force, and lateral force, respectively. When machining along on (010) surface with cutting depth of 1 nm, 2 nm and 3 nm, the calculated cutting forces including tangential, normal, and lateral forces, are indicated in Figure 8. On the initial stage of the cutting process, the tangential and normal forces

start to increase rapidly until the distance of cutting increases to about 10 nm. From then on, Y-27632 2HCl the increasing rate of the cutting force starts to slow down until reaching the steady stage of the cutting process, on which the cutting forces always undulate around the equilibrium value. The lateral force fluctuates around zero because the two side forces of the tool counteract with each other. The fluctuation in cutting force derives from the thermal motion of atoms and the undulation of energy, which results from the deformation of crystal structure during nanometric cutting. Figure 8 Cutting forces. Undeformed chip thickness is (a) 1, (b) 2, and (c) 3 nm. The average tangential and normal forces during the steady stage are calculated when cutting directions are on (010) surface and on (111) surface, respectively.

A previous study has shown that the postmenopausal women in Hong Kong, Beijing and Taiwan have a similar prevalence of morphometric vertebral fracture as Caucasian women in the USA and Europe (about 25% in all regions), in contrast to the marked worldwide variations in the prevalence of hip fractures [21]. The present study further confirmed that, although the risk of hip fractures in Asians was low, Asian men do have a vertebral fracture

risk similar to Caucasian men, and Asian women have an even higher clinical vertebral fracture risk than Caucasian women. The observed ethnic differences in fracture incidences may be due to the fact that hip fracture risk was affected by fall risk, whereas the risk of vertebral fracture mostly depends on bone strength [13]. Despite the low hip fracture rate in our population, Hong Kong women had a higher prevalence BIBW2992 research buy of osteoporosis CFTRinh-172 price (bone mineral density T-score ≤ −2.5 at any one site in reference to ethnic-specific peak young mean according to the ISCD recommendation) than

US Caucasian women (35.8% vs. 20%, respectively) [29, 30] and a similar prevalence of about 6% in Hong Kong and US Caucasian men [31]. In view of the ethnic differences, it is important to obtain accurate information on population fracture risk to characterize the absolute fracture risk of individual subjects. At present, information on the risk of clinical vertebral fracture in Asians is lacking, and the WHO fracture risk assessment algorithms (FRAX®) estimated population-specific absolute major Idasanutlin supplier osteoporotic fracture risks based on the assumption that the ratio of hip-to-vertebral fracture is the same as that observed in Swedish populations to provide. However, our study demonstrated the variations of the spine-to-hip fracture ratios between ethnic groups; thus, a fracture prediction model that assumes a universal spine-to-hip fracture ratio may be biased. Our previous prospective

study on Southern Chinese men over 50 years old has shown that the FRAX® algorithm seemed to overestimate Cepharanthine the 10-year major osteoporotic fracture risk in subjects with low fracture risk, but underestimated the risk for high-risk groups [29]. Results from the current study raise a concern that a model that presumes a ratio of vertebral fractures to hip fractures in a Swedish population might underestimate the risk of vertebral fractures in Asians, resulting in a general underestimation of the absolute risk of major osteoporotic fracture. Strengths of this study include the use of a community-based population to investigate the incidence rate of clinical vertebral fractures. All clinical vertebral fractures and hip fractures were confirmed by the medical record.

Spearman’s LY3009104 cell line correlation analysis indicated a possible relationship between SUV and tumor size in intestinal specimens (rs = 0.50, P < 0.05) (Figure 5a), but not non-intestinal specimens (Figure 5d). The correlation between HK2 or GLUT1 expression and SUV did not find in both cancers (data not shown). There was no correlation between SUV and PCNA mRNA expression in either cancer type (Figure 5b and 5e). Interestingly, the weak association between SUV and HIF1α mRNA expression in intestinal specimens (rs = 0.48, P < 0.05) (Figure 5c) was stronger in non-intestinal specimens

(rs = 0.56, P < 0.01) (Figure 5f). Figure 5 Correlation between mean standardized uptake value and tumor size, hypoxia-inducible factor 1α mRNA levels, or proliferating cell nuclear antigen mRNA levels in intestinal and non-intestinal gastric cancers. (a) Spearman’s correlation analysis indicated a possible RG7112 cell line correlation between standardized uptake value (SUV) and tumor

size in intestinal cancers (rs= 0.50, P < 0.05). (b) No association was found between SUV and proliferating cell nuclear antigen (PCNA) mRNA expression. (c) A weak association was observed between SUV and hypoxia-inducible factor 1α (HIF1α) mRNA expression (rs = 0.48, P < 0.05). (d) In non- intestinal cancer specimens, SUV was not correlated to tumor size. (e) No association was found between SUV and PCNA expression. (f) A significant correlation between SUV and HIF1α mRNA expression was observed (rs = 0.56, P < 0.01). Data are expressed as mean ± SEM. *P < 0.05. HIF1α; Hypoxia-inducible factor 1α, PCNA; Proliferating cell nuclear antigen, SUV; Standardized Uptake Value. Discussion FDG-PET has been used to not only detect cancerous lesions, but also predict therapeutic response after chemotherapy [1, 11, 23]. There are several possible mechanisms behind its ability to reveal malignant potential or cancer cell activity. Our results found that SUV in stage 4 gastric cancer patients was no higher than in stage 2 and stage 3 patients, and the main tumor SUV did not reflect the number of lymph node metastases. Only

tumor size was associated with SUV, a correlation also reported in breast, SCH727965 in vivo pancreatic, and colorectal cancers [20, 24, 25]. These Sitaxentan finding narrow the FDG-PET mechanism possibilities by suggesting that SUV reflects tumor size rather than tumor cell activity for each cancer stage. Over expression of glucose metabolism-related protein in tumors A molecular explanation for high FDG uptake in cancerous tissues is the overexpression of GLUT1, the molecule reported to be responsible for FDG uptake in various cancers [20, 26]. Glucose uptake ability as assessed by FDG-PET was significantly correlated with the doubling time of tumors [27] because increased uptake can provide additional energy to support tumor growth. Yamada et al. [7] determined from immunohistochemistry that GLUT1 expression was an important factor for FDG uptake and also a prognostic tool for gastric cancer. Alakus et al.

Subsequently, phytoene synthase (CrtB) condenses two GGPP molecules yielding the colorless carotenoid phytoene. Four subsequent desaturation reactions by phytoene Selleck AZD8931 desaturase (CrtI) yield the red-colored lycopene [17, 18]. The elongation of lycopene with DMPP to the acyclic C50 carotenoid flavuxanthin is catalyzed by the crtEb gene product lycopene elongase. The cyclization of flavuxanthin to decaprenoxanthin is catalyzed by heterodimeric carotenoid -ɛ-cyclase, encoded by crtY e and crtY f [16, 20, 26]. While mono- and diglucosylated decaprenoxanthin

can be found in C. glutamicum, the genes and enzymes for glucosylation of decaprenoxanthin are still unknown [20]. In this study,

gene-directed GW3965 mw deletion mutagenesis was employed to decipher the functions of the genes present check details in the main carotenogenic gene cluster of C. glutamicum and in a second cluster encoding putative phytoene synthase and phytoene desaturase paralogs. Moreover, the potential of C. glutamicum to produce carotenoids was estimated by metabolic engineering of the conversion of GGPP to lycopene. Results Bioinformatical analysis of the carotenogenic genes The genome of C. glutamicum ATCC 13032 (wild type; WT) encodes genes showing homology to carotenoid biosynthesis genes in two gene clusters that are separated by almost 2 Mbp. The larger cluster is composed of seven genes, crtE (cg0723), cg0722 (encoding a putative membrane protein), crtB (cg0721), crtI (cg0720), crtY e , crtY f (cg01719/18) and crtEb (cg0717) (Figure 1 and 2). The second cluster

consists of a gene putatively encoding phytoene synthase (here named crtB2, cg2672) and two genes with similarity to an N-terminal fragment (crtI2-1, cg2670) and a C-terminal fragment (crtI2-2, cg2668) of phytoene desaturase/dehydrogenase (Figure 1). Figure 1 Genomic organization of the putative and characterized carotenogenic genes in different corynebacteria. Figure 2 Carotenoid biosynthesis in C. glutamicum ATCC 13032 and gene deletion Morin Hydrate and complementation analysis of carotenogenic genes. Cell pellets of C. glutamicum deletion mutants lacking one of the carotenogenic genes crtB, crtI, crtEb or crtY e Y f and the wild type and the corresponding complemented strains (right, EV: empty vector). The cells were grown in 50 ml CGXII medium with 100 mM glucose, inoculated to an OD600 of 1 with a BHI overnight culture. The overexpression was induced at the beginning of the cultivation with 1 mM IPTG. The cluster crtB2/crtI2-1/crtI2-2 has not yet been analyzed. While CrtB and CrtB2 share 49% identity, CrtI2-1 shares 49% identical amino acids with the 364 N-terminal amino acids of CrtI and CrtI2-2 63% identical amino acids with the 104 C-terminal amino acids of CrtI.