To investigate sodium-dependent growth, ‘sodium-free M9’ was prepared by replacing TSA HDAC cell line all sodium salts in M9 (this is around 50 mM Na+ in normal M9) with their potassium equivalents and replacing agar with 0.8% agarose; the sodium-free M9 medium still contained approximately 50 μM sodium from the Amp used for selection. M9 agarose plates required longer incubation

times (up to 4 days) for single colonies to grow. All experiments involving the WT and the ΔnanT strains used cells transformed with the empty vector, i.e. pWKS30. The presence of the vector did not affect the growth phenotypes of either strain (not shown). Starter cultures were prepared as described for the growth experiments, except that o/n growths were carried out in M9 Amp supplemented with 2 mg mL−1 glucose and 1 mM IPTG. Overnight cultures were diluted to an OD650 nm of 0.1 in the

same medium and allowed to grow at 37 °C until they reached an OD650 nm of 0.5, when they were harvested, washed four times in M9, resuspended in the same buffer at a final OD650 nm of 3 and stored www.selleckchem.com/btk.html on ice till use. For Neu5Ac uptake assays, cells were diluted 10-fold in M9 prewarmed at 37 °C and allowed to acclimatize for 2 min, with stirring before initiating the assays by adding of varying amounts of [14C]-Neu5Ac (Sigma) appropriately diluted with unlabelled Neu5Ac. The uptake assay and total protein quantification were then performed as described in Severi et al. (2008), except that 200 μL of cell suspensions were immobilized instead of 400 μL. [14C]-Neu5Ac was normally used at a final concentration of 0.5–2 μM and isotopically diluted (up to 100 ×) with unlabelled Neu5Ac when required. Ks and Vmax values were calculated by fitting the experimental data for uptake rates to a hyperbolic Michaelis–Menton equation using sigmaplot. To assay sodium-dependent Neu5Ac uptake, cells were prepared as for a standard uptake assay, except that sodium-free M9 (see the previous section) was used as both washing and

assay buffers. Salts, i.e. NaCl, KCl and LiCl, were added at a final Methane monooxygenase concentration of 100 mM during the acclimitization phase. The assay was performed as described above with a concentration of 100 μM total Neu5Ac. Cold chase experiments were performed as described in Mulligan et al. (2009), using SEVY1 cells transformed with the appropriate plasmids. To assess the suitability of an E. coliΔnanT strain for the functional characterization of hypothetical Neu5Ac transporters, we first examined cells expressing either nanT itself or the known siaPQM TRAP transporter genes from H. influenzae cloned into a low-copy-number vector under the control of an IPTG-inducible promoter.

Taken together, our results suggest a novel yet unknown

leak K+ channel underlying the pH- and anesthetic-sensitive background conductance in hippocampal astrocytes. “
“Most of us engage in social interactions on a daily basis and the repertoire of social behaviors we acquire during development and later in life are incredibly varied. However, in many neurodevelopmental disorders, including autism spectrum disorders (ASDs), social behavior is severely compromised and indeed this represents a key diagnostic component for such conditions. From genetic association studies, it is increasingly apparent that genes identified as altered in individuals with ASDs often encode synaptic proteins. Fluorouracil cell line Moreover, these synaptic proteins typically serve to scaffold group-I metabotropic glutamate receptors (group-I mGluRs) and ionotropic glutamate receptors (iGluRs; AMPARs and NMDARs), or to enable group-I mGluR to iGluR crosstalk via protein synthesis. Here we aim to explore the possibility of a causal link between altered function of such synaptic proteins and impaired social behaviors that feature in neurodevelopmental disorders, such as ASDs. We review the known synaptic function and role in social behaviors of selected post-synaptic structural proteins (Shank, SAPAP and neuroligin) and regulators of protein

selleck products synthesis (TSC1/2, FMRP and PTEN). While manipulations of proteins involved in group-I mGluR HSP90 and iGluR scaffolding or crosstalk frequently lead to profound alterations in synaptic function and one or more components of social behavior, the neuronal circuits responsible for impairments in specific social behaviors are often poorly defined. We argue for an improved understanding of the neuronal circuits underlying specific social behaviors to aid the development of new ASD therapies. “
“Vision of high temporal resolution depends

on careful regulation of photoresponse kinetics, beginning with the lifetime of activated photopigment. The activity of rhodopsin is quenched by high-affinity binding of arrestin to photoexcited phosphorylated photopigment, which effectively terminates the visual transduction cascade. This regulation mechanism is well established for rod photoreceptors, yet its role for cone vision is still controversial. In this study we therefore analyzed arrestin function in the cone-dominated vision of larval zebrafish. For both rod (arrS ) and cone (arr3 ) arrestin we isolated two paralogs, each expressed in the respective subset of photoreceptors. Labeling with paralog-specific antibodies revealed subfunctionalized expression of Arr3a in M- and L-cones, and Arr3b in S- and UV-cones. The inactivation of arr3a by morpholino knockdown technology resulted in a severe delay in photoresponse recovery which, under bright light conditions, was rate-limiting. Comparison to opsin phosphorylation-deficient animals confirmed the role of cone arrestin in late cone response recovery.

MEP

amplitude at time T after cTBS was defined as the averaged peak-to-peak amplitude of the MEPs recorded during the corresponding batch; this value was then expressed as the change in MEPs compared with pre-cTBS, i.e. [MEPs(T) – MEPs(pre-cTBS)]/MEPs(pre-cTBS). Thus, negative values reflect suppression after cTBS. Student’s t-tests were run to determine if MEP amplitudes were significantly different from zero after cTBS. Bonferroni was applied to correct for multiple comparisons. To account for the variance of the baseline, Student’s t-tests were also run on raw, non-normalized, data. Electroencephalography E7080 in vivo data recorded during batches of single-pulse TMS (Fig. 1C) were processed offline using the EEGlab toolbox (Delorme & Makeig, 2004) running in a MATLAB environment (Mathworks). The EEG signals were analysed with the common reference, as recorded. They were first high-pass filtered above 1 Hz. Continuous data were epoched from 200 ms before the TMS pulse to 600 ms after. Baseline correction was applied based on a pre-TMS interval of 200 ms. Disconnected channels were removed and

recomputed (spherical interpolation) after cleaning (see below). Independent component analysis was performed to separate residual electrical from physiological responses to a TMS pulse. Components related to electrical artifacts were identified by their activity strongly peaking at the vicinity of the stimulation sites during the first tens of milliseconds after a pulse, and by their spectrum covering a restricted frequency range with strong harmonics. Components this website clearly reflecting other artifacts, such as muscle contamination or eye blinks, were also removed. On average, 9.6 ± 4.1 (range 3–17) components were removed, most of the artifacts being identified in the first few components. We cannot exclude that true brain response to TMS was also partly removed with components identified as artifacts. However, as the same components were removed for all conditions within a subject, we

expect changes in EEG response to TMS after cTBS to be related to cTBS-induced changes in brain excitability. Grand-average of TMS-induced for EEG responses were then calculated for the group. For pre-cTBS and for each time batch after cTBS, we calculated the grand-average time-domain response at the C3 electrode (over M1). For each of the pre-cTBS and post-cTBS conditions, we identified the amplitude of four TMS-evoked potentials (TEPs) that are commonly reported in the literature (Paus et al., 2001; Komssi et al., 2004; Bonato et al., 2006; Komssi & Kahkonen, 2006; Van Der Werf & Paus, 2006; Fitzgerald, 2010), i.e. P30, N45, P55 and N100. Then, changes in amplitude compared with pre-cTBS were calculated for each TEP as [TEP(T) – TEP(pre-cTBS)]/TEP(pre-cTBS).

Questionnaires were distributed to the parents to assess awareness of oral health. Results.  There was no significant difference in DMFT scores of study and control group (2.43 +/- 3.72 and 1.36 +/- 2.5 respectively) or in DMFT scores of study and control group (1.5 +/- 1.73 and 1.15 +/- 1.42

respectively), 36% of the study group had untreated caries. Parental knowledge of the link between oral health and infective endocarditis was excellent. Conclusions.  There were no significant differences between the oral health of cardiac children and healthy children although the dmft and DMFT scores of the study group were high. Of concern was the proportion of children with untreated caries in spite of good dental awareness and attendance. “
“Background.  There is only limited

information available in Chile regarding the frequency of biopsied oral lesions in paediatric patients. Aim.  To high throughput screening compounds determine the frequency of histologically diagnosed lesions in oral pathology specimens from paediatric patients in a Chilean population over a 15-year period. Design.  Oral and maxillofacial biopsy records of patients aged 16 years or under http://www.selleckchem.com/products/pirfenidone.html were retrieved by visual inspection from the archives of public and private Oral Pathology Health Services in Valdivia, Chile, during the period 1995–2010. Records that contained anatomical site and histopathological diagnoses of the specimen were included. The study population was divided into three age groups according to dentition

stage. Oral lesions were classified as inflammatory/reactive, cystic, or tumour/tumour-like. Results.  A total of 542 biopsy specimens from children were found. These represented 20.6% of all oral biopsies. The average age was 11.1 years, with female predilection. The most common category of oral lesions was inflammatory/reactive (75.8%), followed by tumour/tumour-like (16.8%) and cystic (7.4%) lesions. The mucocele was the most commonly found lesion, followed by pyogenic granuloma and irritation fibroma, which taken together accounted for 63.8% of all paediatric oral biopsies. The most common localisation for lesions was the lower lip (50.3%). Conclusions.  The vast majority of oral lesions found were predominantly inflammatory/reactive and benign types, although malignant lesions can present themselves in children. “
“International tuclazepam Journal of Paediatric Dentistry 2010; 20: 254–260 Background.  Approximately 10–20% of Streptococcus mutans strains have been reported to possess collagen-binding properties, whereas other species in the oral cavity with those properties remain to be elucidated. Aim.  To identify strains with collagen-binding properties and analyse their characteristics in comparison with S. mutans. Design.  A total of 110 expectorated saliva specimens were collected from 55 pairs of mothers and their children. Bacterial strains with collagen-binding properties were isolated and the species specified.

Almost all primer sets target regions within the 16S rRNA gene with a few exceptions targeting the 16S–23S check details rRNA gene intergenic spacer region and/or the 23S rRNA gene. For simplicity, only the term ‘16S’ is used in the following. The specificity of all primer sets was initially evaluated in silico using nucleotide blast (Altschul et al., 1990) and the Ribosomal Database Project (RDP; Cole et al., 2009). One hundred and ten primer sets found to be suitable after this screening process were synthesized commercially by Eurofins MWG operon GmbH (Ebersberg, Germany). Quantitative real-time PCR was performed on an ABI prism 7900HT

from Applied Biosystems (Nærum, Denmark). All amplification reactions were carried out in transparent 384-well MicroAmp® Optical reaction plates (Applied Biosystems) and sealed with MicroAmp® Selleck AZD2281 Optical Adhesive Film in a total volume of 11 μL containing 5.5 μL 2× SYBR Green PCR Master Mix (Applied Biosystems), 0.4 μL of each primer (10 μM), 2 μL template DNA (2 ng), and 2.7 μL nuclease-free water (Qiagen GmbH, Hilden, Germany). Liquid handling was performed with an epMotion 5075 (Eppendorf, Hørsholm, Denmark). The amplification program was identical for all

amplifications and consisted of one cycle of 50 °C for 2 min; one cycle of 95 °C for 10 min; 40 cycles of 95 °C for 15 s and 60 °C for 1 min; and finally dissociation curve analysis for assessing amplicon specificity (95 °C for 15 s, 60 °C for 15 s, then increasing to 95 °C at 2% ramp rate). Initial qPCR screening on extracted Unoprostone mixed human fecal DNA from healthy volunteers was used in order to identify and remove primer sets, which did not amplify the expected target from this matrix. Fecal DNA was obtained from the control group of a previously conducted study and

was extracted using the QIAamp DNA Stool Mini Kit (Qiagen) preceded by a bead-beater step as previously described (Leser et al., 2000; Licht et al., 2006). A subset of 58 primer sets (of the 110), selected based on their ability to generate amplification products from the complex fecal DNA template material, was used for further evaluation of target specificity on pure culture DNA. The 58 primer sets were tested against extracted DNA from 27 bacterial strains, and one archaeal strain, using the PCR conditions listed above. Reactions were performed in duplicate using 2 ng of DNA as template and always including the universal bacterial primers (reference gene) on the same plate. The generated PCR products were assessed by dissociation curve analysis and 2% agarose gel electrophoresis, stained with SYBR Green, to determine the homogeneity and length of the amplification product, respectively.

Bacterial strains and plasmids used in this study are listed in Supporting Information, Table S1. Photorhabdus luminescens TT01 was grown in Luria–Bertani (LB) medium at 28 °C, and strains of E. coli were grown in LB medium at 37 °C. Escherichia coli DH5α was used as the host Ipilimumab for recombinant DNA cloning. Escherichia coli BL21 (DE3) was used as the host for expression of binary toxin genes. Plasmid pET28a (Novagen) was used as expression vector in BL21 (DE3). Plasmid pETDuet-1 (Novagen) was used as co-expression vector in BL21 (DE3). Total DNA was

extracted from P. luminescens TT01 using the alkali lysis method. It was used as template for amplification of plu1961 and plu1962 (GenBank accession no. BX571865). Oligos used in this study were listed in Table S2. Oligo pair Plu1961-F/Plu1961-R was used to amplify plu1961, and Plu1962-F/Plu1962-R was used to amplify plu1962. Both PCR products were double-digested by EcoRI/SalI and cloned into pET28a to generate plasmids pET-plu1961 and pET-plu1962, respectively. For co-expression of Plu1961 and Plu1962 in BL21 (DE3), Co1961-F/Co1961-R and Co1962-F/Co1962-R were used to amplify plu1961 and plu1962, respectively. PCR products of plu1961 and plu1962 were double-digested

by PstI/SalI and NdeI/XhoI, respectively, and cloned into pETDuet-1 sequentially to generate co-expression plasmid pET-pluBi. All the plasmids were confirmed by DNA sequencing. Plasmids Selleckchem Ganetespib pET-plu1961, pET-plu1962, and pET-pluBi were transformed into BL21 (DE3), and resultant strains were designated as BL21 (plu1961), BL21 (plu1962), and BL (Bi),

respectively. Recombinant strains were grown in LB medium with kanamycin (50 μg mL−1) or ampicillin (100 μg mL−1) at 37 °C to an OD600 between 0.6 and 0.8. Then, isopropyl-beta-d-thiogalactopyranoside (IPTG) was added at a final concentration of 1 mmol L−1. After IPTG induction for 4 h, (-)-p-Bromotetramisole Oxalate aliquots of 1 mL bacteria culture were sampled and harvested by centrifugation (10 000 g, 1 min). Pellets were washed three times with distilled water and suspended in 0.1 mL lysis buffer (50 mmol L−1 NaH2PO4, 300 mmol L−1 NaCl, 10 mmol L−1 imidazole, pH 8.0). Then, cells were lysed by sonication and centrifuged at 10 000 g for 2 min. The supernatant was collected, and 10 μL aliquots were taken for SDS-PAGE. Soluble binary toxins (Plu1961 and Plu1962) were directly purified on 1-mL HisTrapTM HP prepacked columns (GE Healthcare), using an AKTA Purifier system (GE Healthcare; flow rate 1 mL min−1). The column was equilibrated in His A buffer (20 mmol L−1 sodium phosphate, 0.5 mol L−1 NaCl, 20 mmol L−1 imidazole, pH 7.4). Proteins were eluted using a step gradient up to 0.5 mol L−1 imidazole in His A buffer. Fractions were analyzed by SDS-PAGE, and the protein content of the pools was determined using the Bio-Rad Bradford reagent. The purified proteins were dialyzed against PBS buffer prior to application.

, whereas the 162- and 147-bp mpr and zmp products were amplified from B. pseudomallei and B. cepacia, respectively (Fig.

1). All 66 B. pseudomallei, one B. thailandensis and four B. cepacia clinical isolates were positive for the groEL gene, indicating successful detection of the genus Burkholderia. All 65 B. pseudomallei isolates and K96243 strain were positive for the detection of mprA gene. Similarly, all three B. cepacia isolates and ATCC 25416 strain were positive for zmpA gene. Sequence analysis of the PCR products ABT-199 cost from the amplification of groEL, mprA and zmpA matched the published gene sequences in the NCBI website. The negative control strains did not yield any PCR product, suggesting that the primers were highly specific for the different Burkholderia spp. In addition, no cross-reactions were observed within the Burkholderia spp. The mprA and zmpA genes were correctly amplified in the targeted strains, indicating

a specificity of 100%. Akt tumor The limit of detection assay demonstrated that the groEL and zmpA PCR assay was sensitive at 10 pg mL−1 DNA, whereas mprA PCR assay was sensitive at 10 fg mL−1 (Figs 2 and 3). The PCR assay using DNA obtained from blood samples revealed successful amplification of B. pseudomallei in two of the 18 samples tested. On comparison with culture and API 20 NE results, these two PCR-positive samples were also positive for B. pseudomallei by culture and API 20 NE. The PCR-negative samples were also negative on culture, indicating sensitivity and specificity of 100%. However, none of the serum samples produced positive amplicons for any of the three primer sets. Duplex real-time PCR using SYBR green was performed using mprA (162 bp) and zmpA based on the melting curve analysis of amplified products. These primers allowed the amplification of PCR products with distinct melting temperature values, resulting P-type ATPase in the formation of two distinct peaks

representing the two targets. The 167-bp amplicon of mprA (Tm 84 °C) could be clearly separated from the 147-bp amplicon of zmpA (Tm 88 °C) (Figs 4 and 5). No primer dimers were observed in the amplified product, which indicates the specificity of the primers. In this study, a conventional PCR assay was developed for the detection of Burkholderia genus and also for differentiation of the two clinically important human pathogens, B. pseudomallei and B. cepacia. Using bioinformatics tools, this assay incorporated detection of groEL gene, specific for the genus Burkholderia, mprA gene, specific for B. pseudomallei, and zmpA genes specific for B. cepacia. The groEL gene encodes an immunogenic protein of Burkholderia that assists in a proper protein-folding mechanism (Woo et al., 2001). blast analysis revealed that groEL is present in B. mallei, B. pseudomallei, B. cepacia, Burkholderia vietnamiensis and B. thailandensis among the Burkholderiaceae. Moreover, this gene sequence is highly conserved among all Burkholderia spp.

This detection inside the fish cells is not due to the physical disruption of S. parasitica. When the fish cells were treated with recombinant SpHtp1, translocation without the presence of the pathogen

is observed. These results suggest that S. parasitica may have a biotrophic stage in the infection process, which is similar to what has been found in biotrophic and hemibiotrophic plant pathogenic oomycetes. Consequently, it is conceivable that the pathogen has an early infection stage, whereby it does not kill the host cells, but instead, host cells are kept alive in order to enhance its own growth. It is interesting to note that the cells in which SpHtp1 has been translocated, in the presence of S. parasitica (Fig. 2), seem to be somewhat smaller than the surrounding fish cells. It could be that the cells are in fact not smaller, but that the focal plane is not showing the true size of the cells. Erismodegib Alternatively, Saprolegnia is absorbing nutrients from the cells, which results in smaller fish cells. At present, we do not know how many RxLR effectors or whether other RxLR-like effectors are produced

by S. parasitica during an infection. However, the completion of the genome sequence in the near future (at The Broad Institute with Nusbaum, van West, Haas, Russ, Dieguez-Uribeondo and Tyler) will enable a more in-depth analysis of the number of putative RxLR proteins produced by S. parasitica. Talazoparib manufacturer Our work was supported by the BBSRC (I.d.B., K.L.M., A.J.P., S.W., C.J.S., P.v.W.), the University of Aberdeen (E.J.R., V.L.A., C.J.S.,

P.v.W.) and The Royal Society (P.v.W.). We would like to acknowledge the Broad Institute (Carsten Russ, Rays Jiang, Brian Haas and Chad Nusbaum), Brett Tyler (VBI) and P.v.W. for early release of draft supercontigs of the genome sequence of isolate CBS233.65, which helped us choose the best control gene for the Q-PCR experiments and helped resolve the promoter sequence of SpHtp1. I.d.B., K.L.M., A.J.P., E.J.R. and S.W. contributed equally to this work. Fig. S1. Alignment of the full sequences of a subset of oomycete RxLR effector proteins. Fig. S2. Alignment of the upstream region of SpHtp1 with the conserved motif in the oomycete core promoter sequence and genome sequence of SpHtp1. Fig. find more S3. Alignment of SpHtp1 with elicitin-like precursor proteins obtained by blastp analysis of SpHtp1 against nonredundant protein database in NCBI. Fig. S4. Primer sequences used for quantitative real time RT-PCR and reference gene analysis. Fig. S5. SpHtp1 is detected during infection. Fig. S6. Biochemical characterization of SpHtp124-198(His)6. Fig. S7. Uptake of SpHtp1 in fibroblast cells of the rainbow trout cell-line RTG-2. Please note: Wiley-Blackwell is not responsible for the content or functionality of any supporting materials supplied by the authors. Any queries (other than missing material) should be directed to the corresponding author for the article.

Another traveler recalled that she became ill on her friend’s birthday.

Recollection of the symptoms associated with illness episodes appeared to be a more direct task for travelers. All 10 travelers used the calendars provided to recall dates of travel and dates of illness episodes. Four of the 10 travelers used the maps provided to recall the names of the smaller locations. Festival dates provided for each destination country were not used. The final post-travel questionnaires used in the main cohort study were distributed with calendars included. Questionnaires are widely used for data collection. Poorly designed E7080 in vitro questionnaires can affect the quality of data collection, yet there are varying practices in the development and validation of questionnaires. The importance of developing accurate exposure measurements and the impact on the validity of the research conclusions are not well recognized. Furthermore, published papers rarely provide their questionnaires

or reproduce the exact wording of key questions used to define exposures, events, or outcomes. Our objective was to develop and validate a questionnaire for use in a prospective study to estimate the risk of infections in Australian travelers to Asia. Several key features inherent to questionnaires for travelers were Epacadostat cost identified through the development and validation processes. Travelers demonstrated considerable difficulty when attempting to recall the dates surrounding health http://www.selleck.co.jp/products/obeticholic-acid.html events and travel between

major locations. Travel events were recalled in narrative terms and travelers appeared to mentally relive the sequence of events to retrieve the relevant dates from memory. Self-recalled cues or external prompts, such as calendars, provided were used by travelers to formulate a response. Information relating to locations visited or degree of mosquito repellent use was retrieved with less effort than that associated with event dating. Event dating difficulties have been described in a number of other qualitative studies,11 and cognitive research has determined that “when” is the least well-remembered retrieval cue for recalling information about an event from memory.5 Furthermore, there is increased uncertainty about dates with increased time, which is particularly problematic in studies of long-term travel. The diverse experiences of travelers need to be considered when developing items for questionnaires intended for travelers’ study cohorts. This became evident when the response options provided for accommodation types, travel activities, and reasons for travel did not reflect the variety encountered by travelers. In semi-structured interviews, travelers were informative to the expert panel about the breadth of travel experiences, thus contributing to the development of those areas in the questionnaire. There are several recognized methods by which target populations can contribute to the development of questionnaires.

CtpA from P. aeruginosa, however, behaves differently. At least under the experimental conditions used here, this does not contradict our abovementioned hypothesis of an extracellular localization find more of C. trachomatis CtpA, but demonstrates that P. aeruginosa CtpA is in fact a periplasmic protease and that the subcellular localization is an important protein characteristic that must be determined to understand

the physiological role of the protein. The same can be said for CTPs from Gram-positive bacteria as bioinformatic analysis of genomic sequence data suggests that these CTPs are secreted to the extracellular environment. CtpA of P. aeruginosa will remain in the periplasm and is not secreted to the extracellular environment or present in the outer membrane. Several dozen reports have been published about bacterial CTPs after the initial study of Hara et Bortezomib ic50 al. (1991). Most refer to CTPs as periplasmic proteases, although the experimental evidence

for the individual protein was not given. As far as we know, we are the first to confirm experimentally the exclusive periplasmic localization of a CTP-3 of P. aeruginosa. The periplasmic localization of CtpA strictly excludes the possibility that the protein is directly involved in the virulence of P. aeruginosa as an extracellular effector molecule. The obvious role of CTPs in the virulence of pathogenic bacteria, as shown experimentally in B. suis, B. bacilliformis and B. mallei (Mitchell & Minnick, 1997; Bandara et al., 2005, 2008; Lad et al., 2007) and P. aeruginosa (R. Hoge et al., unpublished data), must be due to an indirect effect mediated by those a substrate protein of CTP in the context of a periplasmic function. Equivalent to the evolution and function of CTPs from phototrophic organisms, CTPs from Gram-negative

bacteria may be required to activate periplasmatic proteins by cleavage, just as the photosynthetic D1 protein is activated in plant cells. A good candidate as a substrate protein would be the PBP-3. Their periplasmic localization would support evidence of the Prc substrates in E. coli identified by their subcellular localization, because PBP-3 is anchored in the cytoplasmic membrane with the C-terminal end facing the periplasm (Nguyen-Distèche et al., 1998). As PBP-3 in E. coli is involved in the essential process of cell wall synthesis and the CTP could function as an activator of PBP-3, E. coli PBP-3 is thought to be a key element cell division in which it presumably initiates polymerization of the septum peptidoglycan by catalysing a transpeptidation reaction during cell division (Nguyen-Distèche et al., 1998).