ABSTRACT
Hybrid molecules were constructed with either polyclonal antibodies against Trypanosoma cruzi antigens or monoclonal antibody against Trypanosoma brucei brucei low-density lipoprotein (LDL)-receptor conjugated with chlorambucil. Physical-chemical analysis of the hybrid molecule showed four chlorambucil coupling sites in each IgG and a binding constant in the order of 10(4). Maintenance of IgG integrity was indicated by its circular dichroism pattern. Biologic activity of the hybrid molecule was shown by its inhibitory effect on the mobility and proliferation of the parasite. An IgG-chlorambucil conjugate, produced with monoclonal antibody anti-T. b. brucei LDL-receptor, led to the immobilization of the T. cruzi forms, albeit at a much lesser level than that obtained with a mouse polyclonal anti-T. cruzi IgG linked to the drug. Targeting experimental T. cruzi infection with a specific IgG-chlorambucil conjugate resulted in consistent reduction of parasitemia and mortality, thus showing its potential usefulness in controlling the acute form of the disease.
Subject(s)
Chagas Disease/therapy , Chlorambucil/therapeutic use , Immunoconjugates/therapeutic use , Immunoglobulin G/therapeutic use , Animals , Cell Membrane/metabolism , Chagas Disease/parasitology , Chagas Disease/pathology , Immunoconjugates/metabolism , Mice , Mice, Inbred BALB C , Movement/physiology , Parasitemia/therapy , Rabbits , Survival Analysis , Trypanosoma cruzi/growth & development , Trypanosoma cruzi/metabolism , Trypanosoma cruzi/physiologyABSTRACT
The cytolytic seed protein enterolobin from seeds of Enterolobium contortisiliquum was purified by using FPLC on a Mono Q column giving a single peak in capillary electrophoresis. The complete amino acid sequence of the plant cytolysin was determined by an automated method, yielding a molecular mass of 54,806 Da. Databank searches and sequence alignment demonstrated a high degree of sequence identity and similarity between enterolobin and bacterial aerolysins from Aeromonas hydrophila and A. sobria. Several key residues involved in oligomerization of A. hydrophila aerolysin are conserved in enterolobin. Circular dichroism measurements and structural predictions revealed that enterolobin is very rich in beta sheet, like aerolysin. Light-scattering studies revealed that enterolobin oligomerizes as a hexamer at pH levels below 7.0. NaCl concentrations above 50 mM caused dimerization of enterolobin. Dithiothreitol did not cause oligomerization.
Subject(s)
Cytotoxins/chemistry , Plant Proteins/chemistry , Trees/chemistry , Amino Acid Sequence , Amino Acids/analysis , Bacterial Toxins/chemistry , Electrophoresis, Capillary , Light , Molecular Sequence Data , Pore Forming Cytotoxic Proteins , Protein Precursors/chemistry , Protein Structure, Secondary , Scattering, Radiation , Seeds/chemistry , Sequence Analysis , Sequence Homology, Amino AcidABSTRACT
1) The presence of histamine in prickles and lead and stem tissues of Cnidosculus oligandrus (Mull. Arg) Pax & Hoffm (Euphorbiaceae) is reported. 2) Prickles on stems and leaves have larger amounts of histamine, as determined by the isolated guinea-pig ileum and by fluorimetry of the condensation product of the histamine and ortho-phthalaldehyde (OPT). 3) Additionally, thin-layer chromatography and the increase of vascular permeability bioassay were used to confirm the presence of histamine.
Subject(s)
Histamine/isolation & purification , Plant Extracts/analysis , Animals , Capillary Permeability/drug effects , Chromatography, Thin Layer , Diphenhydramine/pharmacology , Female , Fluorometry , Guinea Pigs , Ileum/physiology , Male , Muscle Contraction/drug effects , Spectrometry, FluorescenceABSTRACT
Os espectros de absorcao de segunda derivada e de derivada-de-diferenca do inibidor triptico e quimotriptico de sementes de Vigna unguiculata (L) Walp. (var. Serido) foram obtidos para determinar o estado dos residuos de fenilalanina na proteina.O espectro de segunda derivada do inibidor nativo, comparado aquele do inibidor reduzido e carboximetilado e ao de uma mistura-modelo de N-acetil etil esteres de fenilalanina, tirosina e triptofanio (em relacao molar de 3:1:1, para imitar a proteina ana), sugere que os residuos de fenilalanina sao extensamente expostos ao solvente. Alem disso, a primeira derivada dos espectros de diferenca do inibidor nativo e do reduzido indica que os tres residuos de fenilalanina por molecula de inibidor nativo estao expostos ao solvente em grau aproximadamente igual ao da proteina reduzida e carboximetilada
