ABSTRACT
N-Hydroxyformamide-class metalloprotease inhibitors were designed and synthesized, which have potent broad-spectrum activity versus matrix metalloproteases and TNF-alpha converting enzyme (TACE). Compound 13c possesses good oral and intravenous pharmacokinetics in the rat and dog.
Subject(s)
Enzyme Inhibitors/pharmacology , Formamides/pharmacology , Matrix Metalloproteinase Inhibitors , Metalloendopeptidases/antagonists & inhibitors , Oligopeptides/pharmacology , ADAM Proteins , ADAM17 Protein , Animals , Dogs , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacokinetics , Formamides/chemistry , Oligopeptides/chemistry , Rats , Structure-Activity RelationshipABSTRACT
Rabbit antibodies to purified human placental galactose-1-phosphate uridyltransferase (EC 2.7.7.12) were used to establish immunologic cross-reactivity patterns for the enzyme in hemolysates, prepared from red cells of a normal individual, a homozygous Duarte variant, and a heterozygous Los Angeles variant. The antibody immunoprecipitated all three forms of the enzyme. The amount of antibody absorbed by each hemolysate was related to the different levels of activity, and examination of hemolysate/antibody reaction mixtures by starch gel electrophoresis revealed that the antibody quantitatively precipitated all of the isoenzyme forms that characterize these three genetic variants.
Subject(s)
Galactosemias/enzymology , Isoenzymes/analysis , Nucleotidyltransferases/analysis , UTP-Hexose-1-Phosphate Uridylyltransferase/analysis , Animals , Cells, Cultured , Cross Reactions , Electrophoresis, Starch Gel , Erythrocytes/enzymology , Fibroblasts/enzymology , Galactosemias/genetics , Heterozygote , Homozygote , Humans , Isoenzymes/genetics , Precipitin Tests , Rabbits , UTP-Hexose-1-Phosphate Uridylyltransferase/geneticsABSTRACT
Red blood cell lysates from normal individuals, a homozygous Duarte variant, and a patient with transferase-deficiency galactosemia were challenged with rabbit antibody to pure human placental galactose-1-phosphate uridylyltransferase. Although the antibody quantitatively precipitated the enzymatically active proteins in the normal and Duarte hemolysates, the Duarte sample absorbed only about one-half as much antibody as did the normal. In contrast, the antibody did not react with the galactosemic hemolysate.
Subject(s)
Erythrocytes/enzymology , Galactosemias/enzymology , Transferases/deficiency , Adult , Animals , Antibodies/isolation & purification , Humans , Immunochemistry , Immunosorbent Techniques , Male , Rabbits , Transferases/bloodABSTRACT
Isolated liver nucleoli from rats treated for 3 days with thioacetamide contained an enzyme activity which specifically degraded conjugate protein A24. Two-dimensional polyacrylamide gel electrophoresis indicated that the amount of protein A24 in chromatin decreased during incubation at 37 degrees C for 60 min with these nucleoli. Concomitantly, a marked increase was found in the content of free ubiquitin, the nonhistone component of protein A24. Incubation of 3H-labeled protein A24 with the thioacetamide-treated liver nucleoli resulted in the linear release of 3H-labeled histone 2A and 3H-free ubiquitin in the presence of phenylmethanesulfonyl fluoride (PMSF) for 2 h. Pretreatment of the nucleoli with trypsin or by heating at 80 degrees C for 10 min inhibited their ability to cleave protein A24. Protein A24 lyase catalyzes the reaction: protein A24 leads to histone 2A plus ubiquitin.
