Protein stability, flexibility and function

Proteins rely on flexibility to respond to environmental changes, ligand binding and chemical modifications. Potentially, a perturbation that changes the flexibility of a protein may interfere with its function. Millions of mutations have been performed on thousands of proteins in quests for a delineation of the molecular details of their function. Several of these mutations interfered with the binding of a specific ligand with a concomitant effect on the stability of the protein scaffold. It has been ambiguous and not straightforward to recognize if any relationships exist between the stability of a protein and the affinity for its ligand. In this review, we present examples of proteins where changes in stability results in changes in affinity and of proteins where stability and affinity are uncorrelated. We discuss the possibility for a relationship between stability and binding. From the data presented is it clear that there are specific sites (flexibility hotspots) in proteins that are important for both binding and stability. This article is part of a Special Issue entitled: Protein Dynamics: Experimental and Computational Approaches.     

Maternal Use of Antibiotics and the Risk of Childhood Febrile Seizures: A Danish Population-Based Cohort

Objective

In a large population-based cohort in Denmark to examine if maternal use of antibiotics during pregnancy, as a marker of infection, increases the risk of febrile seizures in childhood in a large population-based cohort in Denmark.

Methods

All live-born singletons born in Denmark between January 1, 1996 and September 25, 2004 and who were alive on the 90^th day of life were identified from the Danish National Birth Registry. Diagnoses of febrile seizures were obtained from the Danish National Hospital Register and maternal use of antibiotics was obtained from the National Register of Medicinal Product Statistics. Hazard ratios (HR) and 95% confidence intervals (95% CI) were estimated by Cox proportional hazard regression models.

Results

We followed 551,518 singletons for up to 5 years and identified a total of 21,779 children with a diagnosis of febrile seizures. Slightly increased hazard ratios were observed among most exposure groups when compared to the unexposed group, ex. HR 1.08 95% CI: 1.05–1.11 for use of any systemic antibiotic during pregnancy.

Conclusion

We found weak associations between the use of pharmacologically different antibiotics during pregnancy and febrile seizures in early childhood which may indicate that some infections, or causes or effects of infections, during pregnancy could affect the fetal brain and induce susceptibility to febrile seizures.     

Individual Variation in Influenza A Virus Infection Histories and Long-Term Immune Responses in Mallards

Wild dabbling ducks (genus Anas) are the main reservoir for influenza A virus (IAV) in the Northern Hemisphere. Current understanding of disease dynamics and epidemiology in this virus-host system has primarily been based on population-level surveillance studies and infection experiments conducted in laboratory settings. Using a combined experimental-natural approach with wild-strain captive mallards (Anas platyrhynchos), we monitored individual IAV infection histories and immunological responses of 10 birds over the course of 15 months. This is the first detailed study to track natural IAV infection histories over several seasons amongst the same individuals growing from juvenile to adults. The general trends in the infection histories of the monitored birds reflected seasonal variation in prevalence at the population level. However, within the study group there were significant differences between individuals in infection frequency as well as in short and long term anti-IAV antibody response. Further observations included individual variation in the number of infecting virus subtypes, and a strong tendency for long-lasting hemagglutinin-related homosubtypic immunity. Specifically, all infections in the second autumn, except one, were of different subtypes compared to the first autumn. The variation among birds concerning these epidemiologically important traits illustrates the necessity for IAV studies to move from the level of populations to examine individuals in order to further our understanding of IAV disease and epidemiology.     

Extracellular Matrix Aggregates from Differentiating Embryoid Bodies as a Scaffold to Support ESC Proliferation and Differentiation

Embryonic stem cells (ESCs) have emerged as potential cell sources for tissue engineering and regeneration owing to its virtually unlimited replicative capacity and the potential to differentiate into a variety of cell types. Current differentiation strategies primarily involve various growth factor/inducer/repressor concoctions with less emphasis on the substrate. Developing biomaterials to promote stem cell proliferation and differentiation could aid in the realization of this goal. Extracellular matrix (ECM) components are important physiological regulators, and can provide cues to direct ESC expansion and differentiation. ECM undergoes constant remodeling with surrounding cells to accommodate specific developmental event. In this study, using ESC derived aggregates called embryoid bodies (EB) as a model, we characterized the biological nature of ECM in EB after exposure to different treatments: spontaneously differentiated and retinoic acid treated (denoted as SPT and RA, respectively). Next, we extracted this treatment-specific ECM by detergent decellularization methods (Triton X-100, DOC and SDS are compared). The resulting EB ECM scaffolds were seeded with undifferentiated ESCs using a novel cell seeding strategy, and the behavior of ESCs was studied. Our results showed that the optimized protocol efficiently removes cells while retaining crucial ECM and biochemical components. Decellularized ECM from SPT EB gave rise to a more favorable microenvironment for promoting ESC attachment, proliferation, and early differentiation, compared to native EB and decellularized ECM from RA EB. These findings suggest that various treatment conditions allow the formulation of unique ESC-ECM derived scaffolds to enhance ESC bioactivities, including proliferation and differentiation for tissue regeneration applications.     

Development and Validation of a Vitamin D Status Prediction Model in Danish Pregnant Women: A Study of the Danish National Birth Cohort

Vitamin D has been hypothesized to reduce risk of pregnancy complications such as preeclampsia, gestational diabetes mellitus, and preterm delivery. However, many of these outcomes are rare and require a large sample size to study, representing a challenge for cohorts with a limited number of preserved samples. The aims of this study were to (1) identify predictors of serum 25-hydroxy-vitamin D (25(OH)D) among pregnant women in a subsample (N = 1494) of the Danish National Birth Cohort (DNBC) and (2) develop and validate a score predicting 25(OH)D-status in order to explore associations between vitamin D and maternal and offspring health outcomes in the DNBC. In our study sample, 42.3% of the population had deficient levels of vitamin D (<50 nmol/L 25(OH)D) and average levels of 25(OH)D-status were 56.7(s.d. 24.6) nmol/L. A prediction model consisting of intake of vitamin D from diet and supplements, outdoor physical activity, tanning bed use, smoking, and month of blood draw explained 40.1% of the variance in 25(OH)D and mean measured 25(OH)D-level increased linearly by decile of predicted 25(OH)D-score. In total 32.2% of the women were placed in the same quintile by both measured and predicted 25(OH)D-values and 69.9% were placed in the same or adjacent quintile by both methods. Cohen''s weighted kappa coefficient (Κ = 0.3) reflected fair agreement between measured 25(OH)D-levels and predicted 25(OH)D-score. These results are comparable to other settings in which vitamin D scores have shown similar associations with disease outcomes as measured 25(OH)D-levels. Our findings suggest that predicted 25(OH)D-scores may be a useful alternative to measured 25(OH)D for examining associations between vitamin D and disease outcomes in the DNBC cohort, but cannot substitute for measured 25(OH)D-levels for estimates of prevalence.     

Climate change impact on seaweed meadow distribution in the North Atlantic rocky intertidal

The North-Atlantic has warmed faster than all other ocean basins and climate change scenarios predict sea surface temperature isotherms to shift up to 600 km northwards by the end of the 21st century. The pole-ward shift has already begun for many temperate seaweed species that are important intertidal foundation species. We asked the question: Where will climate change have the greatest impact on three foundational, macroalgal species that occur along North-Atlantic shores: Fucus serratus, Fucus vesiculosus, and Ascophyllum nodosum? To predict distributional changes of these key species under three IPCC (Intergovernmental Panel on Climate Change) climate change scenarios (A2, A1B, and B1) over the coming two centuries, we generated Ecological Niche Models with the program MAXENT. Model predictions suggest that these three species will shift northwards as an assemblage or “unit” and that phytogeographic changes will be most pronounced in the southern Arctic and the southern temperate provinces. Our models predict that Arctic shores in Canada, Greenland, and Spitsbergen will become suitable for all three species by 2100. Shores south of 45° North will become unsuitable for at least two of the three focal species on both the Northwest- and Northeast-Atlantic coasts by 2200. If these foundational species are unable to adapt to the rising temperatures, they will lose their centers of genetic diversity and their loss will trigger an unpredictable shift in the North-Atlantic intertidal ecosystem.     

Campylobacter jejuni Actively Invades the Amoeba Acanthamoeba polyphaga and Survives within Non Digestive Vacuoles

The Gram-negative bacterium Campylobacter jejuni is able to enter, survive and multiply within the free living amoeba Acanthamoeba polyphaga, but the molecular mechanisms behind these events are still unclear. We have studied the uptake and intracellular trafficking of viable and heat killed bacterial cells of the C. jejuni strain 81–176 in A. polyphaga. We found that viable bacteria associated with a substantially higher proportion of Acanthamoeba trophozoites than heat killed bacteria. Furthermore, the kinetics of internalization, the total number of internalized bacteria as well as the intracellular localization of internalized C. jejuni were dramatically influenced by bacterial viability. Viable bacteria were internalized at a high rate already after 1 h of co-incubation and were observed in small vacuoles tightly surrounding the bacteria. In contrast, internalization of heat killed C. jejuni was low at early time points and did not peak until 96 h. These cells were gathered in large spacious vacuoles that were part of the degradative pathway as determined by the uptake of fluorescently labeled dextran. The amount of heat killed bacteria internalized by A. polyphaga did never reach the maximal amount of internalized viable bacteria. These results suggest that the uptake and intracellular survival of C. jejuni in A. polyphaga is bacterially induced.     

Computed Tomography Measurement of Rib Cage Morphometry in Emphysema

Background

Factors determining the shape of the human rib cage are not completely understood. We aimed to quantify the contribution of anthropometric and COPD-related changes to rib cage variability in adult cigarette smokers.

Methods

Rib cage diameters and areas (calculated from the inner surface of the rib cage) in 816 smokers with or without COPD, were evaluated at three anatomical levels using computed tomography (CT). CTs were analyzed with software, which allows quantification of total emphysema (emphysema%). The relationship between rib cage measurements and anthropometric factors, lung function indices, and %emphysema were tested using linear regression models.

Results

A model that included gender, age, BMI, emphysema%, forced expiratory volume in one second (FEV₁)%, and forced vital capacity (FVC)% fit best with the rib cage measurements (R² = 64% for the rib cage area variation at the lower anatomical level). Gender had the biggest impact on rib cage diameter and area (105.3 cm²; 95% CI: 111.7 to 98.8 for male lower area). Emphysema% was responsible for an increase in size of upper and middle CT areas (up to 5.4 cm²; 95% CI: 3.0 to 7.8 for an emphysema increase of 5%). Lower rib cage areas decreased as FVC% decreased (5.1 cm²; 95% CI: 2.5 to 7.6 for 10 percentage points of FVC variation).

Conclusions

This study demonstrates that simple CT measurements can predict rib cage morphometric variability and also highlight relationships between rib cage morphometry and emphysema.     

Congenital Cerebral Palsy,Child Sex and Parent Cardiovascular Risk

Objective

Genes associated with cardiovascular disease may also be risk factors for congenital cerebral palsy (CP) and these associations may be modified by sex, since there is an increased risk of CP in male children. We investigated the association between CP of the child with cardiovascular disease in parents, taking sex of the child into consideration.

Methods

All parents of non-adopted singletons born in Denmark between 1973 and 2003 were included. Parents of a child with CP, confirmed by the Danish National CP registry, were considered exposed. Cox proportional hazards regressions were used to model risk of cardiovascular outcomes for exposed parents compared to all other parents beginning at the child’s 10^th birthday.

Results

We identified 733,730 mothers and 666,652 fathers among whom 1,592 and 1,484, respectively, had a child with CP. The mean age for mothers at end of follow up was 50±8 years. After adjustment for maternal age, parental education, child’s sex, child’s residence, child being small for gestational age and maternal hypertensive disorder during pregnancy, mothers of CP male children had an excess risk of cardiovascular disease (HR: 1.52, 95% CI: 1.16-2.00), attributable mostly to an increased incidence of hypertension and cerebrovascular disease. After additional adjustment for preterm birth, the association was markedly attenuated for cardiovascular disease (1.34, 95%CI: 1.02 - 1.76), became nonsignificant for hypertension, but remained significant for cerebrovascular disease (HR: 2.73, 95% CI: 1.45- 5.12). There was no increased risk of cardiovascular events in mothers of female CP children, or fathers of CP children of any sex.

Conclusions

Women that have a male child with CP are at increased risk for premature cardiovascular disease. Part of this association may be related to risk factors for preterm births.