Functional insights from proteome-wide structural modeling of <Emphasis Type="Italic">Treponema pallidum</Emphasis> subspecies <Emphasis Type="Italic">pallidum</Emphasis>, the causative agent of syphilis

Background

Syphilis continues to be a major global health threat with 11 million new infections each year, and a global burden of 36 million cases. The causative agent of syphilis, Treponema pallidum subspecies pallidum, is a highly virulent bacterium, however the molecular mechanisms underlying T. pallidum pathogenesis remain to be definitively identified. This is due to the fact that T. pallidum is currently uncultivatable, inherently fragile and thus difficult to work with, and phylogenetically distinct with no conventional virulence factor homologs found in other pathogens. In fact, approximately 30% of its predicted protein-coding genes have no known orthologs or assigned functions. Here we employed a structural bioinformatics approach using Phyre2-based tertiary structure modeling to improve our understanding of T. pallidum protein function on a proteome-wide scale.

Results

Phyre2-based tertiary structure modeling generated high-confidence predictions for 80% of the T. pallidum proteome (780/978 predicted proteins). Tertiary structure modeling also inferred the same function as primary structure-based annotations from genome sequencing pipelines for 525/605 proteins (87%), which represents 54% (525/978) of all T. pallidum proteins. Of the 175 T. pallidum proteins modeled with high confidence that were not assigned functions in the previously annotated published proteome, 167 (95%) were able to be assigned predicted functions. Twenty-one of the 175 hypothetical proteins modeled with high confidence were also predicted to exhibit significant structural similarity with proteins experimentally confirmed to be required for virulence in other pathogens.

Conclusions

Phyre2-based structural modeling is a powerful bioinformatics tool that has provided insight into the potential structure and function of the majority of T. pallidum proteins and helped validate the primary structure-based annotation of more than 50% of all T. pallidum proteins with high confidence. This work represents the first T. pallidum proteome-wide structural modeling study and is one of few studies to apply this approach for the functional annotation of a whole proteome.

     

Dose effects of modified alternate-day fasting regimens on in vivo cell proliferation and plasma insulin-like growth factor-1 in mice.

Reduced cell proliferation is associated with lower cancer risk. Alternate-day fasting (ADF), defined as alternating 24-h periods of ad libitum feeding and fasting, decreases cell proliferation. The effect of modified regimens of ADF on cell proliferation, however, has not been examined. This study measured the effects of modified ADF regimens on prostate and splenic T-cell proliferation and circulating insulin-like growth factor-1 (IGF-1) levels in mice. In a 4-wk study, 24 male C57BL/6J mice were randomized to one of four interventions: 1) ADF-25% [25% calorie restriction (CR) on fast day], 2) ADF-50% (50% CR on fast day), 3) ADF-100% (100% CR on fast day), and 4) control. Body weight of the ADF-100% group was less (P < 0.005) than that of the ADF-25% and ADF-50% groups posttreatment. On the feast day, the ADF-100% and ADF-50% groups ate 85% and 45% more food, respectively, than controls, indicating a hyperphagic response to fasting. Proliferation rates of T-cells were 6% and 30% lower (P < 0.05) in the ADF-50% and ADF-100% groups, respectively, relative to controls. Prostate cell proliferation was reduced (P < 0.05) by 49% in the ADF-100% group, relative to controls, but did not change in the other groups. IGF-1 levels were reduced (P < 0.05) by 40%, relative to controls, in the ADF-100% group. These findings confirm the beneficial effects of ADF-100% on cancer risk by decreasing cell proliferation and IGF-1 levels and suggest that modified ADF regimens comprising 25-50% CR on the fast day do not replicate these effects.     

On the presence and localization of glycogen accumulations inside coronet cells of the saccus vasculosus of the dogfish (Scylliorhinus caniculus)

Summary In several coronet cells of the saccus vasculosus of Scylliorhinus large quantities of glycogen occur, as shown by light and electron microscopy. The significance of glycogen as an energy storage necessary for a transcellular ion transport process taking place in the coronet cells is discussed.The authors thank Dr. F.C.G. van de Veerdonk, W. F. Jansen and W. F. G. Flight for reading the manuscipt and for their critical remarks. They are also indebted to Mr. H. van Kooten and his staff for their valuable photographic assistance.     

Permafrost thaw and intense thermokarst activity decreases abundance of stream benthic macroinvertebrates

Intensification of permafrost thaw has increased the frequency and magnitude of large permafrost slope disturbances (mega slumps) in glaciated terrain of northwestern Canada. Individual thermokarst disturbances up to 40 ha in area have made large volumes of previously frozen sediments available for leaching and transport to adjacent streams, significantly increasing sediment and solute loads in these systems. To test the effects of this climate‐sensitive disturbance regime on the ecology of Arctic streams, we explored the relationship between physical and chemical variables and benthic macroinvertebrate communities in disturbed and undisturbed stream reaches in the Peel Plateau, Northwest Territories, Canada. Highly disturbed and undisturbed stream reaches differed with respect to taxonomic composition and invertebrate abundance. Minimally disturbed reaches were not differentiated by these variables but rather were distributed along a disturbance gradient between highly disturbed and undisturbed sites. In particular, there was evidence of a strong negative relationship between macroinvertebrate abundance and total suspended solids, and a positive relationship between abundance and the distance from the disturbance. Increases in both sediments and nutrients appear to be the proximate cause of community differences in highly disturbed streams. Declines in macroinvertebrate abundance in response to slump activity have implications for the food webs of these systems, potentially leading to negative impacts on higher trophic levels, such as fish. Furthermore, the disturbance impacts on stream health can be expected to intensify as climate change increases the frequency and magnitude of thermokarst.     

Insulin Stimulates S100B Secretion and These Proteins Antagonistically Modulate Brain Glucose Metabolism

Neurochemical Research - Brain metabolism is highly dependent on glucose, which is derived from the blood circulation and metabolized by the astrocytes and other neural cells via several pathways....     

The role of soil conditions in fine root ecomorphology in Norway spruce (Picea abies (L.) Karst.)

The present study is an attempt to investigate the pattern of morphological variability of the short roots of Norway spruce (Picea abies (L.) Karst.) growing in different soils. Five root parameters – diameter, length and dry weight of the root tip, root density (dry weight per water-saturated volume) and specific root area (absorbing area of dry weight unit) were studied with respect to 11 soil characteristics using CANOCO RDA analysis. The investigation was conducted in seven study areas in Estonia differing in site quality class and soil type. Ten root samples per study area were collected randomly from the forest floor and from the 20 cm soil surface layer. Eleven soil parameters were included in the study: humus content, specific soil surface area, field capacity, soil bulk density, pH (KCl and H₂O dilution's), N and Ca concentrations, Ca/Al and C/N ratios, and the decomposition rate of fine roots (<2 mm dia.). Root morphological characteristics most strongly related to the measured soil characteristics in the different sites were specific root area, root density and diameter of the short roots, the means varying from 29 to 42 m² kg⁻¹, from 310 to 540 kg m⁻³ and from 0.26 to 0.32 mm, respectively; root density being most sensitive. The most favourable site and soil types resulting in fine roots with morphological characteristics for optimizing nutrient uptake (e.g. low short root density and high specific root area) were Umbric Luvisol (Oxalis), Dystric Gleysol (Oxalis) and Gleyic Luvisol (Hepatica). These soil types correspond to highly productive natural forest stands of Norway spruce in Estonia. All measured soil variables explained 28% of total variance of the root characteristics. The most important variables related to root morphology were the humus content, field capacity and specific soil surface area. This revised version was published online in June 2006 with corrections to the Cover Date.     

Pentachlorophenol Dechlorination in Fluidized-Bed Reactors under Methanogenic Conditions

The reductive dechlorination of chlorophenols was studied in three fluidized-bed reactors (FBRs) with respect to enrichment, pathways, complete dechlorination, and overall performance. The methanogenic consortia, developed by previous researchers in our laboratory, have been further enriched by reducing the ratio of substrate to pentachlorophenol (PCP) and increasing the PCP loading. The performance of the consortia was improved, and complete dechlorination at high PCP loading rates was observed, reaching a PCP loading of 1227 µmol/L d with 99% chlorophenol removal. The dechlorination rates in the reactors for chlorophenol (CP) congeners were obtained and were used to evaluate the performance of the three consortia and to quantitatively estimate the fates of these chlorophenols in the reactors. The consortium with the best performance was further investigated in bottle tests by treatment with heat and metabolic inhibitors to examine chlorophenol degradation and to characterize the CP degraders. The degradation of all monochlorophenols was completely inhibited after heat treatment, but the degradation of all other tested chlorophenols was hardly affected by heat treatment, indicating that spore-forming bacteria likely were involved in dechlorination. Addition of sulfate negatively affected CP degradation, but addition of molybdate reduced the effect of sulfate. Tests with 2-bromoethanesulfonic acid and vancomycin indicated that bacteria were responsible for chlorophenol degradation in the consortium.     

Acute Effects of Pathogenic Simian-Human Immunodeficiency Virus Challenge on Vaccine-Induced Cellular and Humoral Immune Responses to Gag in Rhesus Macaques

Simian-human immunodeficiency virus (SHIV) infection in macaques provides a convenient model for testing vaccine efficacy and for understanding viral pathogenesis in AIDS. We immunized macaques with recombinant, Salmonella typhimurium (expressing Gag) or soluble Gag in adjuvant to generate T-cell-dependent lymphoproliferative or serum antibody responses. Immunized animals were challenged by intrarectal inoculation with SHIV89.6PD. Virus infection was accompanied by rapid losses of lymphoproliferative responses to Gag or phytohemagglutinin. By 8 weeks, mitogen responses recovered to near normal levels but antigen-specific immunity remained at low or undetectable levels. Serum antibody levels were elevated initially by virus exposure but soon dropped well below levels achieved by immunization. Our studies show a rapid depletion of preexisting Gag-specific CD4⁺ T cells that prevent or limit subsequent antiviral cellular and humoral immune responses during acute SHIV infection.     

Quantitative characterization of the mechanism of action and impact of a ‘proteolysis-permitting’ anti-PCSK9 antibody

A recent report described a novel mechanism of action for an anti-proprotein convertase subtilisin-kexin type 9 (PCSK9) monoclonal antibody (LY3015014, or LY), wherein the antibody has improved potency and duration of action due to the PCSK9 epitope for LY binding. Unlike other antibodies, proteolysis of PCSK9 can occur when LY is bound to PCSK9. We hypothesized that this allowance of PCSK9 cleavage potentially improves LY efficiency through two pathways, namely lack of accumulation of intact PCSK9 and reduced clearance of LY. A quantitative modeling approach is necessary to further understand this novel mechanism of action. We developed a mechanism-based model to characterize the relationship between antibody pharmacokinetics, PCSK9 and LDL cholesterol levels in animals, and used the model to better understand the underlying drivers for the improved efficiency of LY. Simulations suggested that the allowance of cleavage of PCSK9 resulting in a lack of accumulation of intact PCSK9 is the major driver of the improved potency and durability of LY. The modeling reveals that this novel ‘proteolysis-permitting’ mechanism of LY is a means by which an efficient antibody can be developed with a total antibody dosing rate that is lower than the target production rate. We expect this engineering approach may be applicable to other targets and that the mathematical models presented herein will be useful in evaluating similar approaches.     

A rapid method for determining dynamic binding capacity of resins for the purification of proteins

Biolayer interferometry is a novel method for quantifying macromolecules, such as proteins, in solution. The presence of other, non-binding molecules does not interfere with quantification, which allows one to measure the concentration of the molecule of interest in a crude mixture. Here we apply this method to determining the dynamic binding capacity of affinity resins.