Nuclear magnetic resonance structure shows that the severe acute respiratory syndrome coronavirus-unique domain contains a macrodomain fold

The nuclear magnetic resonance (NMR) structure of a central segment of the previously annotated severe acute respiratory syndrome (SARS)-unique domain (SUD-M, for "middle of the SARS-unique domain") in SARS coronavirus (SARS-CoV) nonstructural protein 3 (nsp3) has been determined. SUD-M(513-651) exhibits a macrodomain fold containing the nsp3 residues 528 to 648, and there is a flexibly extended N-terminal tail with the residues 513 to 527 and a C-terminal flexible tail of residues 649 to 651. As a follow-up to this initial result, we also solved the structure of a construct representing only the globular domain of residues 527 to 651 [SUD-M(527-651)]. NMR chemical shift perturbation experiments showed that SUD-M(527-651) binds single-stranded poly(A) and identified the contact area with this RNA on the protein surface, and electrophoretic mobility shift assays then confirmed that SUD-M has higher affinity for purine bases than for pyrimidine bases. In a further search for clues to the function, we found that SUD-M(527-651) has the closest three-dimensional structure homology with another domain of nsp3, the ADP-ribose-1 ''-phosphatase nsp3b, although the two proteins share only 5% sequence identity in the homologous sequence regions. SUD-M(527-651) also shows three-dimensional structure homology with several helicases and nucleoside triphosphate-binding proteins, but it does not contain the motifs of catalytic residues found in these structural homologues. The combined results from NMR screening of potential substrates and the structure-based homology studies now form a basis for more focused investigations on the role of the SARS-unique domain in viral infection.

NMR structure shows that the SARS-unique domain contains a macrodomain fold

The NMR structure of a central segment of the previously annotated "SARS-unique domain" (SUD-M; "middle of the SARS-unique domain") in the SARS coronavirus (SARS-CoV) non-structural protein 3 (nsp3) has been determined. SUD-M(513-651) exhibits a macrodomain fold containing the nsp3-residues 528-648, and there is a flexibly extended N-terminal tail with the residues 513-527 and a C-terminal flexible tail of residues 649-651. As a follow-up to this initial result, we also solved the structure of a construct representing only the globular domain of residues 527-651 [SUD-M(527-651)]. NMR chemical shift perturbation experiments showed that SUD-M(527-651) binds single-stranded poly-A and identified the contact area with this RNA on the protein surface, and electrophoretic mobility shift assays then confirmed that SUD-M has higher affinity for purine bases than for pyrimidine bases. In further search for clues to the function, we found that SUD-M(527-651) has the closest three-dimensional structure homology with another domain of nsp3, the ADP-ribose-1''-phosphatase nsp3b, although the two proteins share only 5% sequence identity in the homologous sequence regions. SUD-M(527-651) also shows 3D structure homology with several helicases and NTP-binding proteins, but it does not contain the motifs of catalytic residues found in these structural homologues. The combined results from NMR screening of potential substrates and the structure-based homology studies now form a basis for more focused investigations on the role of the SARS-unique domain in viral infection.

SARS coronavirus unique domain: three-domain molecular architecture in solution and RNA binding

Nonstructural protein 3 of the severe acute respiratory syndrome (SARS) coronavirus includes a "SARS-unique domain" (SUD) consisting of three globular domains separated by short linker peptide segments. This work reports NMR structure determinations of the C-terminal domain (SUD-C) and a two-domain construct (SUD-MC) containing the middle domain (SUD-M) and the C-terminal domain, and NMR data on the conformational states of the N-terminal domain (SUD-N) and the SUD-NM two-domain construct. Both SUD-N and SUD-NM are monomeric and globular in solution; in SUD-NM, there is high mobility in the two-residue interdomain linking sequence, with no preferred relative orientation of the two domains. SUD-C adopts a frataxin like fold and has structural similarity to DNA-binding domains of DNA-modifying enzymes. The structures of both SUD-M (previously determined) and SUD-C (from the present study) are maintained in SUD-MC, where the two domains are flexibly linked. Gel-shift experiments showed that both SUD-C and SUD-MC bind to single-stranded RNA and recognize purine bases more strongly than pyrimidine bases, whereby SUD-MC binds to a more restricted set of purine-containing RNA sequences than SUD-M. NMR chemical shift perturbation experiments with observations of (15)N-labeled proteins further resulted in delineation of RNA binding sites (i.e., in SUD-M, a positively charged surface area with a pronounced cavity, and in SUD-C, several residues of an anti-parallel beta-sheet). Overall, the present data provide evidence for molecular mechanisms involving the concerted actions of SUD-M and SUD-C, which result in specific RNA binding that might be unique to the SUD and, thus, to the SARS coronavirus.

Configurations institutionnelles de la protection de l’enfance: regards croisés de l’Afrique, de l’Europe et de l’Amérique du Nord

Cet article porte sur l’analyse de trois configurations institutionnelles de la protection de l’enfance : celle en place au Burkina Faso, en Belgique et au Québec. Pour chaque configuration, le texte explore les transformations qui ont marqué le passage de la prise en charge exclusive de l’enfant par la famille vers la présence accrue de l’État et la manière dont la Convention internationale des droits de l’enfant de 1989 a influencé ce passage. Il montre, au travers d’une lecture historique, que l’implication de l’État dans la protection des enfants a connu des formes et des modalités variables selon le temps et l’espace. L’approche actuelle en matière de protection de l’enfance au Nord et au Sud, approche largement inspirée de la perspective des droits, représente un défi tant pour les intervenants que pour les familles, car son application dépend à fois des ressources disponibles pour aider les enfants et les familles en difficulté, de la capacité d’action des institutions publiques et de l’efficacité des interventions. This article deals with the analysis of three institutional configurations of child protection: those in Burkina Faso, in Belgium, and in Québec. With respect to each configuration, the text explores the changes from situations in which the family had sole control of the child to those where the State played a greater role, and the manner in which the 1989 International Convention on the Rights of the Child has affected such changes. It shows, through a reading of history, that the involvement of the State in child protection has gone through different forms and stages over time and space. The current strategies as regards child protection in the North and in the South—an approach that is largely subject to a legal perspective—represent a challenge both for intervenors and for families, since their application depends on the resources available for helping children and families in difficulty, on the ability of the public institutions to intervene, and on the efficiency of such interventions.

Mobilisations contre le travail des enfants au Burkina Faso : quelquees considérations de la dimension genre

La lutte contre le travail des enfants au Burkina Faso a resurgi dans la scène publique ces dix dernières années avec un engouement sans précédent aussi bien de la part des acteurs étatiques, des ONG que des organismes onusiens. Ce chapitre traite, dans une double perspective sociologique et anthropologique, de la dimension « genre » de cette lutte et cherche à voir dans quelle mesure celle-ci est prise en compte (ou non) dans les pratiques concrètes de retrait d’enfants des travaux considérés comme dangereux. Peut-on parler de politiques et de programmes « genrés » dans le champ de la lutte contre le travail des enfants au Burkina Faso ? Le chapitre est structuré en trois parties. Il présente d’abord les éléments caractéristiques de l’approche genre dans la lutte contre le travail des enfants. Ensuite, il aborde les politiques et les acteurs de cette lutte au Burkina Faso, en décrivant les processus de problématisation et publicisation de la question. Enfin, il traite du cas spécifique d’un projet triennal de retrait d’enfants des mines artisanales des régions du Nord et du Sahel burkinabè pour questionner la prise en compte du genre, des obstacles sociaux, culturels et d’autres contraintes possibles à l’égalité fille-garçon dans cette intervention. Il questionne par ailleurs la part des acteurs de terrain dans la (re)production des normes et des rôles sexués. Les données sont issues de quatre années de recherche menée au Burkina Faso entre 2008 et 2011, auprès d’acteurs engagés dans la production des politiques de lutte et/ou engagés dans la lutte concrète, mais également avec des enfants, parents et employeurs.