The two tryptophans of β2-microglobulin have distinct roles in function and folding and might represent two independent responses to evolutionary pressure

We have recently discovered that the two tryptophans of human β2-microglobulin have distinctive roles within the structure and function of the protein. Deeply buried in the core, Trp95 is essential for folding stability, whereas Trp60, which is solvent-exposed, plays a crucial role in promoting the binding of β2-microglobulin to the heavy chain of the class I major histocompatibility complex (MHCI). We have previously shown that the thermodynamic disadvantage of having Trp60 exposed on the surface is counter-balanced by the perfect fit between it and a cavity within the MHCI heavy chain that contributes significantly to the functional stabilization of the MHCI. Therefore, based on the peculiar differences of the two tryptophans, we have analysed the evolution of β2-microglobulin with respect to these residues

Biological passport parameters

For long time anti-doping authorities have tried to find a way to tackle the use of forbidden substances and methods (namely erythropoiesis stimulating agents and blood transfusion) which improve sport performance through an increase of red cells and therefore oxygen transfer. From the end of the ‘90s scientists have explored two different but complimentary ways to detect these substances/methods: direct detection, aiming to find the forbidden agent in an athlete’s biological sample and indirect detection, through the measurement of hematological parameters which are modified by blood doping. The biological passport is the most recent and sophisticated tool in regard to indirect doping detection. Its principles rely on the monitoring of relevant biomarkers on a regular basis so as to constitute an individual and longitudinal profile for any given athlete, with the subject becoming his/her own reference. Standardized procedures in relation to blood sampling, transport and analysis have to be respected in order to decrease pre-analytical and analytical variability. A statistical adaptive model is applied to interpret all the gathered data. The evaluation of this information follows forensic principle, where multiple piece of evidence can be added in order to support the opinion that a doping offence has taken place. Finally, the biological passport experience of the International Cycling Union is presented

Time for change: a roadmap to guide the implementation of the World Anti-Doping Code 2015

A medical and scientific multidisciplinary consensus meeting was held from 29 to 30 November 2013 on Anti-Doping in Sport at the Home of FIFA in Zurich, Switzerland, to create a roadmap for the implementation of the 2015 World Anti-Doping Code. The consensus statement and accompanying papers set out the priorities for the antidoping community in research, science and medicine. The participants achieved consensus on a strategy for the implementation of the 2015 World Anti-Doping Code. Key components of this strategy include: (1) sport-specific risk assessment, (2) prevalence measurement, (3) sport-specific test distribution plans, (4) storage and reanalysis, (5) analytical challenges, (6) forensic intelligence, (7) psychological approach to optimise the most deterrent effect, (8) the Athlete Biological Passport (ABP) and confounding factors, (9) data management system (Anti-Doping Administration & Management System (ADAMS), (10) education, (11) research needs and necessary advances, (12) inadvertent doping and (13) management and ethics: biological data. True implementation of the 2015 World Anti-Doping Code will depend largely on the ability to align thinking around these core concepts and strategies. FIFA, jointly with all other engaged International Federations of sports (Ifs), the International Olympic Committee (IOC) and World Anti-Doping Agency (WADA), are ideally placed to lead transformational change with the unwavering support of the wider antidoping community. The outcome of the consensus meeting was the creation of the ad hoc Working Group charged with the responsibility of moving this agenda forward

Time for change: a roadmap to guide the implementation of the World Anti-Doping Code 2015

A medical and scientific multidisciplinary consensus meeting was held from 29 to 30 November 2013 on Anti-Doping in Sport at the Home of FIFA in Zurich, Switzerland, to create a roadmap for the implementation of the 2015 World Anti-Doping Code. The consensus statement and accompanying papers set out the priorities for the antidoping community in research, science and medicine. The participants achieved consensus on a strategy for the implementation of the 2015 World Anti-Doping Code. Key components of this strategy include: (1) sport-specific risk assessment, (2) prevalence measurement, (3) sport-specific test distribution plans, (4) storage and reanalysis, (5) analytical challenges, (6) forensic intelligence, (7) psychological approach to optimise the most deterrent effect, (8) the Athlete Biological Passport (ABP) and confounding factors, (9) data management system (Anti-Doping Administration & Management System (ADAMS), (10) education, (11) research needs and necessary advances, (12) inadvertent doping and (13) management and ethics: biological data. True implementation of the 2015 World Anti-Doping Code will depend largely on the ability to align thinking around these core concepts and strategies. FIFA, jointly with all other engaged International Federations of sports (Ifs), the International Olympic Committee (IOC) and World Anti-Doping Agency (WADA), are ideally placed to lead transformational change with the unwavering support of the wider antidoping community. The outcome of the consensus meeting was the creation of the ad hoc Working Group charged with the responsibility of moving this agenda forward

The New Apolipoprotein A-I Variant Leu(174) → Ser Causes Hereditary Cardiac Amyloidosis, and the Amyloid Fibrils Are Constituted by the 93-Residue N-Terminal Polypeptide

We identified a novel missense mutation in the apolipoprotein A-I gene, T2069C Leu(174) → Ser, in a patient affected by familial systemic nonneuropathic amyloidosis. The amyloid deposits mostly affected the heart of the proband, who underwent transplantation for end-stage congestive heart failure. Amyloid fibrils of myocardial and periumbilical fat samples immunoreacted exclusively with anti-ApoA-I antibodies. Amyloid fibrils extracted from the heart were constituted, according to amino acid sequencing and mass spectrometry analysis, by an amino-terminal polypeptide ending at Val(93) of apolipoprotein A-I (apoA-I); no other significant fragments were detected. The mutation segregates with the disease; it was demonstrated in the proband and in an affected uncle and excluded in three healthy siblings. The plasma levels of high-density lipoprotein and apoA-I were significantly lower in the patient than in unaffected individuals. This represents the first case of familial apoA-I amyloidosis in which the mutation is outside the polypeptide fragment deposited as fibrils. Visualization of the mutation in the three-dimensional structure of lipid-free apoA-I, composed of four identical polypeptide chains, indicates that position 174 of one chain is located near position 93 of an adjacent chain and suggests that the amino acid replacement in position 174 is permissive for a proteolytic split at the C-terminal of Val(93)

Effect of tetracycline on β₂-m induced locomotory defect in transgenic <i>C. elegans</i> strains.

<p>Egg-synchronized control worms (vector), wild type β₂-m expressing worms (WT), P32G-mutated β₂-m and ΔN6-truncated β₂-m expressing nematodes (ΔN6) were placed at 20°C into fresh NMG plates seeded with tetracycline-resistant <i>E. coli</i>. At their L3/L4 larval stage, animals were fed with 50–100 µM tetracycline hydrochloride or 100 µM doxycycline (100 µl/plate). Body bends in liquid were scored after 24 hours. At least three independent assays were performed. Data are mean of number of body bends/min ± SD; **p<0.01 vs. the Vector, °°p<0.01 vs. the respective untreated group, according to one-way ANOVA (N = 60 animals for each group).</p

Behavioural phenotypes of transgenic <i>C. elegans</i> strains.

<p>(<b>A</b>) Larval growth of control worms (Vector), wild type β₂-m expressing worms (WT) and nematodes expressing P32G or 7–99 truncated form of β₂-m (ΔN6). One hundred synchronized eggs were placed into fresh NMG plates seeded with OP50 as food, and the number of L1/L2, L2/L3 and L4/adult worms were scored after 24, 48 and 72 hours, respectively. Data are expressed as percentage of total worms in the plate at each time point and are given as mean of three independent experiments (N = 300). (<b>B</b>) Correlation between oligomers of β₂-m and reduction in growth rate of transgenic <i>C. elegans</i> strains. Percentage of adult worms of each transgenic strain, scored 72 after egg synchronization, was correlated to the the amount of A11-positive oligomeric assemblies detected by dot blotting. Data of both graphic axes represent mean of three independent experiments. (<b>C</b>) Kaplan-Meier survival curves of transgenic hermaphrodite adult nematodes. Animals were placed in plates seeded with OP50 starting from L4, cultured at 20°C and transferred to fresh plates for each consecutive other days. Survival rate was scored every day and expressed as percent of survival. Plots are representative of three independent experiments (N = 30). (<b>D</b>) Body bends in liquid of transgenic worms. At least three independent assays were performed (N = 100 animals for each group). Data are given as mean of number of body bends/min ± SE, *p<0.05 and **p<0.01 vs. the vector, °°p<0.01 vs. WT, according to one-way ANOVA. (<b>E</b>) Superoxide anions production in control worms (Vector), wild type β₂-m expressing worms (WT) and in nematodes expressing P32G or 7–99 truncated form of β₂-m (ΔN6). Age-synchronized worms were collected in PBS containing 1.6 ml of 1% Tween 20 and colorimetric NBT assay was carried out as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0052314#s2" target="_blank">Materials and Methods</a>. Results show the fold increase in superoxide production calculated as NBT absorbance/mg of proteins (% NBT) compared to Vector; *p<0.05 vs. vehicle and ° p<0.05 vs. WT, according to one-way ANOVA. Error bars indicate SD.</p