The N-Terminal residues 43 to 60 form the interface for dopamine mediated α-synuclein dimerisation

α-synuclein (α-syn) is a major component of the intracellular inclusions called Lewy bodies, which are a key pathological feature in the brains of Parkinson's disease patients. The neurotransmitter dopamine (DA) inhibits the fibrillisation of α-syn into amyloid, and promotes α-syn aggregation into SDS-stable soluble oligomers. While this inhibition of amyloid formation requires the oxidation of both DA and the methionines in α-syn, the molecular basis for these processes is still unclear. This study sought to define the protein sequences required for the generation of oligomers. We tested N- (α-syn residues 43-140) and C-terminally (1-95) truncated α-syn, and found that similar to full-length protein both truncated species formed soluble DA: α-syn oligomers, albeit 1-95 had a different profile. Using nuclear magnetic resonance (NMR), and the N-terminally truncated α-syn 43-140 protein, we analysed the structural characteristics of the DA:α-syn 43-140 dimer and α-syn 43-140 monomer and found the dimerisation interface encompassed residues 43 to 60. Narrowing the interface to this small region will help define the mechanism by which DA mediates the formation of SDS-stable soluble DA:α-syn oligomers

Intramolecular Cohesion of Coils Mediated by Phenylalanine–Glycine Motifs in the Natively Unfolded Domain of a Nucleoporin

The nuclear pore complex (NPC) provides the sole aqueous conduit for macromolecular exchange between the nucleus and the cytoplasm of cells. Its diffusion conduit contains a size-selective gate formed by a family of NPC proteins that feature large, natively unfolded domains with phenylalanine–glycine repeats (FG domains). These domains of nucleoporins play key roles in establishing the NPC permeability barrier, but little is known about their dynamic structure. Here we used molecular modeling and biophysical techniques to characterize the dynamic ensemble of structures of a representative FG domain from the yeast nucleoporin Nup116. The results showed that its FG motifs function as intramolecular cohesion elements that impart order to the FG domain and compact its ensemble of structures into native premolten globular configurations. At the NPC, the FG motifs of nucleoporins may exert this cohesive effect intermolecularly as well as intramolecularly to form a malleable yet cohesive quaternary structure composed of highly flexible polypeptide chains. Dynamic shifts in the equilibrium or competition between intra- and intermolecular FG motif interactions could facilitate the rapid and reversible structural transitions at the NPC conduit needed to accommodate passing karyopherin–cargo complexes of various shapes and sizes while simultaneously maintaining a size-selective gate against protein diffusion

Raynaud's syndrome in children : systematic review and development of recommendations for assessment and monitoring

Objective. To develop recommendations for investigation and monitoring of children with Raynaud's syndrome, based on paediatric evidence collated by a systematic review. Method. A systematic review was undertaken to establish the paediatric evidence for assessment and monitoring of Raynaud's syndrome. An expert panel including members of the Paediatric Rheumatology European Society (PRES) Scleroderma Working Group, were invited to a consensus meeting where recommendations were developed based on evidence graded by the systematic review and where evidence was lacking, consensus opinion. A nominal technique was used where 75% consensus was taken as agreement. Results. The expert panel recommended testing anti-nuclear antibody (ANA), more specific antibodies associated with connective tissue disease and nail-fold capillaroscopy in all children presenting with Raynaud's syndrome as data suggests these can be risk factors for evolution into a connective tissue disease. The frequency of follow-up recommended depends on presence of these risk factors with the aim to detect evolving connective tissue disease early in high risk individuals. Those with no abnormalities on capillaroscopy and negative autoantibodies were deemed low risk of progression, whereas those with ANA positivity, specific autoantibodies and/or nailfold capillary changes were deemed high risk and more frequent follow-up was recommended. Conclusion. Recommendations, primarily based on consensus opinion, were agreed regarding investigation and monitoring of children who present with Raynaud's syndrome. Further prospective studies are needed to better define the risk factors for progression to connective tissue disease

Raynaud's syndrome in children: systematic review and development of recommendations for assessment and monitoring.

Objective. To develop recommendations for investigation and monitoring of children with Raynaud’s syndrome, based on paediatric evidence collated by a systematic review. Method. A systematic review was undertaken to establish the paediatric evidence for assessment and monitoring of Raynaud’s syndrome. An expert panel including members of the Paediatric Rheumatology European Society (PRES) Scleroderma Working Group, were invited to a consensus meeting where recommendations were developed based on evidence graded by the systematic review and where evidence was lacking, consensus opinion. A nominal technique was used where 75% consensus was taken as agreement. Results. The expert panel recommended testing anti-nuclear antibody (ANA), more specific antibodies associated with connective tissue disease and nail-fold capillaroscopy in all children presenting with Raynaud’s syndrome as data suggests these can be risk factors for evolution into a connective tissue disease. The frequency of follow-up recommended depends on presence of these risk factors with the aim to detect evolving connective tissue disease early in high risk individuals. Those with no abnormalities on capillaroscopy and negative autoantibodies were deemed low risk of progression, whereas those with ANA positivity, specific autoantibodies and/or nailfold capillary changes were deemed high risk and more frequent follow-up was recommended. Conclusion. Recommendations, primarily based on consensus opinion, were agreed regarding investigation and monitoring of children who present with Raynaud’s syndrome. Further prospective studies are needed to better define the risk factors for progression to connective tissue disease

Raynaud's syndrome in children: systematic review and development of recommendations for assessment and monitoring

Objective. To develop recommendations for investigation and monitoring of children with Raynaud's syndrome, based on paediatric evidence collated by a systematic review. Method. A systematic review was undertaken to establish the paediatric evidence for assessment and monitoring of Raynaud's syndrome. An expert panel including members of the Paediatric Rheumatology European Society (PRES) Scleroderma Working Group, were invited to a consensus meeting where recommendations were developed based on evidence graded by the systematic review and where evidence was lacking, consensus opinion. A nominal technique was used where 75% consensus was taken as agreement. Results. The expert panel recommended testing anti-nuclear antibody (ANA), more specific antibodies associated with connective tissue disease and nail-fold capillaroscopy in all children presenting with Raynaud's syndrome as data suggests these can be risk factors for evolution into a connective tissue disease. The frequency of follow-up recommended depends on presence of these risk factors with the aim to detect evolving connective tissue disease early in high risk individuals. Those with no abnormalities on capillaroscopy and negative autoantibodies were deemed low risk of progression, whereas those with ANA positivity, specific autoantibodies and/or nailfold capillary changes were deemed high risk and more frequent follow-up was recommended. Conclusion. Recommendations, primarily based on consensus opinion, were agreed regarding investigation and monitoring of children who present with Raynaud's syndrome. Further prospective studies are needed to better define the risk factors for progression to connective tissue disease.Alder Hey Childrens NHS Fdn Trust, Liverpool, Merseyside, EnglandSemmelweis Univ, Budapest, HungaryUniv Childrens Hosp Ljubljana, Ljubljana, SloveniaUniv Childrens Hosp, Tubingen, GermanyUniv Klinikum Hamburg Eppendorf, Hamburg, Germany[Piotto, D. P.] Univ Fed Sao Paulo, Sao Paulo, BrazilIstanbul Kanuni Sultan Suleyman Educ & Res Hosp, Istanbul, TurkeyCharles Univ Prague, Prague, Czech RepublicCath Childrens Hosp, Dept Paediat Dermatol, Hamburg, GermanyUniv Genoa, Genoa, ItalyGhent Univ Hosp, Ghent, BelgiumHamburger Zentrum Kinder & Jugendrheumatol, Hamburg, GermanyUniv Fed Sao Paulo, Sao Paulo, BrazilWeb of Scienc

A specific nanobody prevents amyloidogenesis of D76N β2-microglobulin in vitro and modifies its tissue distribution in vivo

Systemic amyloidosis is caused by misfolding and aggregation of globular proteins in vivo for which effective treatments are urgently needed. Inhibition of protein self-aggregation represents an attractive therapeutic strategy. Studies on the amyloidogenic variant of β2-microglobulin, D76N, causing hereditary systemic amyloidosis, have become particularly relevant since fibrils are formed in vitro in physiologically relevant conditions. Here we compare the potency of two previously described inhibitors of wild type β2-microglobulin fibrillogenesis, doxycycline and single domain antibodies (nanobodies). The β2-microglobulin -binding nanobody, Nb24, more potently inhibits D76N β2-microglobulin fibrillogenesis than doxycycline with complete abrogation of fibril formation. In β2-microglobulin knock out mice, the D76N β2-microglobulin/ Nb24 pre-formed complex, is cleared from the circulation at the same rate as the uncomplexed protein; however, the analysis of tissue distribution reveals that the interaction with the antibody reduces the concentration of the variant protein in the heart but does not modify the tissue distribution of wild type β2-microglobulin. These findings strongly support the potential therapeutic use of this antibody in the treatment of systemic amyloidosis