Monoclonal antibody mapping of the envelope glycoprotein of the dengue 2 virus, Jamaica

Although dengue (DEN) virus is the etiologic agent of dengue fever, the most prevalent vector-borne viral disease in the world, precise information on the antigenic structure of the dengue virion is limited. We have prepared a set of murine monoclonal antibodies (Mabs) specific for the envelope (E) glycoprotein of DEN 2 virus and used these antibodies in a comprehensive biological and biochemical analysis to identify 16 epitopes. Following domain nomenclature developed for the related flavivirus, tick-bourne encephalitis, three functional domains were identified. Five epitopes associated with domain A were arranged in three spatially independently regions. These A-domain epitopes were destroyed by reduction, and antibodies reactive with these epitopes were able to block virus hemagglutination, neutralize virus infectivity, and block virus haemagglutination, neutralize virus infectivity, and block virus-mediated cell membrane fusion. Domain-A epitopes were present on the full-length E glycoprotein, a 45-kDa tryptic peptide representing its first 400 amino acids (aa) and a 22-kDA tryptic peptide representing at least aa 1-120. Four epitopes mapped into domain B, as determined by their partial resistance to reduction and the localization of these epitopes on a 9-kDa tryptic or chymotryptic peptide fragment (aa 300-400). One domain-B-reactive MAb was also capable of binding to a DEN 2 synthetic peptide corresponding to aa 333-351 of the E glycoprotein, confirming the location of this domain. Domain-B epitopes elicited MAbs that were potent neutralizers of virus infectivity and blocked hemagglutination, but they did not block virus-mediated cell-membrane fusion. Domains A and B were spatially associated. As with tick-bourne encephalitis virus, determination of domain C was more problematic: however, at least four epitopes and biochemical characteristics consistent with C-domain epitopes(AU)

Cytochrome c oxidase: a new conformational model

Cytochrome c oxidase and zinc cytochrome c were cross-linked to form a 1:1 enzyme-substrate convalent complex. The reaction was catalyzed by 1-ethyl-3-[3-(dimethylamino) propyl] carbodiimide (EDC) and cytochrome c, the substrate, became attached to the binding site located at submit II of the enzyme. This covalent complex could still be reduced by unbound ferrocytochrome c, suggesting the existence of more than one substrate binding site [1]. Previously [2] we reported that the reduction of heme a triggered a major conformational change within cytochrome c oxidase. Here we propose a model which incorporates these two findings with other established features of cytochrome c oxidase. The most prominent features of this model are (i) Electrons may enter the enzyme via both heme and CUa. Thus cytochrome oxidas may be regarded as having two substrate binding sites, one near heme a and the other near CUa. (ii) At any instant however, only one site is normally available for the binding of substrate. (iii) Subsequent to binding, a substrate molecule transfers an electron to the redox center at the site; reduction of the redox centre triggers a conformational transition which abolishes that binding site and simultaneously leads to the formation of an adjacent substrate binding site near the other redox centre. (iv) The two sites possess approximately equivalent affinity for substrate(AU)

Hemoglobin O Arab in four negro families and its interaction with hemoglobin S and hemoglobin C

Hemoglobin O Arab (O2á2 121 Glu leads to Lys) was found in 25 members of four apparently unrelated Negro families in the West Indian island of Jamaica. In each family the propositus had Hb SO disease. Two cases had been mistakenly diagnosed as Hb SC disease. Two persons heterozygous for both Hb C and Hb O Arab were found in these families, and Hb O Arab á thalassemia in one other relative. The clinical course and symptomatology of Hb SO disease is comparable to that in Hb SD(O2á2 121 Glu leads to GluNH2) disease and more severe than Hb SC disease. In vitro mixtures of Hb O Arab and Hb S change from a liquid to a gel phase at total hemoglobin concentrations weaker than those required to gel pure Hb S, whereas mixtures of Hb C require a stronger total hemoglobin concentration before gelling will occur. Oxygen dissociation studies on red cells containing Hb SO show a homozygous sickle-cell anemia and outside the range for subjects with sickle-cell Hb C disease.(AU)