RESUMO
Proteoglycans (PGs), including heparan sulfate forms, are important regulators of tumor progression. In the PGs biosynthetic process, the core protein is synthesized on a ribosomal template and the sugar chains are assembled post-translationally, one sugar at a time, starting with the linkage of xylose to a serine residue of the core protein and followed by galactosidation of the xylosylprotein. Hydrophobic xylopyranosides have been previously shown to prime heparan sulfate synthesis, a property that was required to cause growth inhibition of tumor cells. To know if the antiproliferative activity of synthetic xylopyranosides is related to their ability to act as "decoy acceptors" of xylosylprotein 4-ß-galactosyltransferase, we have heterologously expressed the catalytic domain of the human ß-1,4-GalT 7 and studied the ability of a variety of synthetic xylopyranoside derivatives to act as substrates or inhibitors of the recombinant enzyme.
Assuntos
Galactosiltransferases , Glicopeptídeos , Glicosídeos , N-Acetil-Lactosamina Sintase/metabolismo , Naftóis , Proteínas Recombinantes/metabolismo , Sequência de Aminoácidos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/metabolismo , Galactosiltransferases/síntese química , Galactosiltransferases/metabolismo , Galactosiltransferases/farmacologia , Regulação Bacteriana da Expressão Gênica , Glicopeptídeos/síntese química , Glicopeptídeos/metabolismo , Glicopeptídeos/farmacologia , Glicosídeos/síntese química , Glicosídeos/metabolismo , Glicosídeos/farmacologia , Humanos , Dados de Sequência Molecular , N-Acetil-Lactosamina Sintase/genética , Naftóis/síntese química , Naftóis/metabolismo , Naftóis/farmacologia , Proteínas Recombinantes/genética , SolubilidadeAssuntos
Galactosiltransferases/metabolismo , Proteínas Recombinantes/metabolismo , Uridina Difosfato N-Acetilgalactosamina/síntese química , Uridina Difosfato N-Acetilglicosamina/síntese química , Técnicas de Química Combinatória , Proteínas de Escherichia coli/metabolismo , Galactosiltransferases/síntese química , Galactosiltransferases/química , Humanos , Estrutura Molecular , Complexos Multienzimáticos/metabolismo , Proteínas Recombinantes/síntese química , Proteínas Recombinantes/química , Especificidade por Substrato , Uridina Difosfato N-Acetilgalactosamina/química , Uridina Difosfato N-Acetilgalactosamina/metabolismo , Uridina Difosfato N-Acetilglicosamina/química , Uridina Difosfato N-Acetilglicosamina/metabolismoRESUMO
Sandwich capture enzyme immunoassays (EIA I and EIA II) are described. Water soluble B substance was reacted simultaneously with affinity-purified dinitrophenyl goat anti-B IgG and affinity-purified goat anti-B IgG-peroxidase conjugate. The complex formed of B substance, dinitrophenyl IgG and IgG-peroxidase conjugate was trapped onto a polystyrene ball coated with affinity-purified goat anti-dinitrophenyl bovine serum albumin IgG. After washing, peroxidase activity bound to the ball was assayed by fluorometry (the sandwich capture EIA I). In the sandwich capture EIA II, the complex was, after thorough washing, eluted from the ball with dinitrophenyl-L-lysine, and then peroxidase activity in the eluate was assayed. The thorough washing and elution processes improved the sensitivity 3.3-fold, and B substance in saliva samples from type B and AB secretors could be detected 200- to 500-fold more sensitively than hemagglutination inhibition, a method commonly used in forensic practices.
