RESUMO
1-(N-Phenyl)amino-1-deoxy-α-D-manno-hept-2-ulose (2) and two multivalent BSA-based structures 7 and 8, d-manno-configured C-glycosyl-type compounds derived from an Amadori rearrangement, were evaluated as ligands for mannoside-specific lectins of various sources. The determination of the concentration corresponding to 50% of inhibition (IC50) is described. Multivalency turned out to effectively influence ligand selectivity and lectin binding.
Assuntos
Antibacterianos/farmacologia , Lectinas/farmacologia , Manosídeos/farmacologia , Amaryllidaceae/efeitos dos fármacos , Antibacterianos/química , Burkholderia/efeitos dos fármacos , Canavalia/efeitos dos fármacos , Galanthus/efeitos dos fármacos , Lectinas/síntese química , Lectinas/química , Ligantes , Manosídeos/química , Testes de Sensibilidade Microbiana , Estrutura Molecular , Vicia/efeitos dos fármacosRESUMO
Bauhinia purpurea lectin (BPA) is one of the beta-galactose-binding leguminous lectins. Leguminous lectins contain a long metal-binding loop, part of which determines their carbohydrate-binding specificities. Random mutations were introduced into a portion of the cDNA coding BPA that corresponds to the carbohydrate-binding loop of the lectin. An library of the mutant lectin expressed on the surface of lambda foo phages was screened by the panning method. Several phage clones with an affinity for mannose or N-acetylglucosamine were isolated. These results indicate the possibility of making artificial lectins (so-called "cyborg lectins") with distinct and desired carbohydrate-binding specificities.
Assuntos
Fabaceae/genética , Lectinas/síntese química , Lectinas/genética , Proteínas de Plantas/síntese química , Proteínas de Plantas/genética , Plantas Medicinais , Acetilglucosamina/análogos & derivados , Acetilglucosamina/genética , Acetilglucosamina/metabolismo , Sequência de Aminoácidos , Bacteriófago lambda/genética , Bacteriófago lambda/metabolismo , Sítios de Ligação/genética , Fabaceae/química , Vetores Genéticos/síntese química , Lectinas/biossíntese , Lectinas/metabolismo , Manose/genética , Manose/metabolismo , Proteínas de Membrana/biossíntese , Proteínas de Membrana/genética , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Biblioteca de Peptídeos , Lectinas de Plantas , Proteínas de Plantas/biossíntese , Proteínas de Plantas/metabolismo , Engenharia de Proteínas , Albumina Sérica/genética , Albumina Sérica/metabolismo , Soroalbumina Bovina/genética , Soroalbumina Bovina/metabolismo , Vírion/genética , Vírion/metabolismoRESUMO
Monoclonal antibodies (monAbs) displaying diverse affinity to native and recombinant mistletoe lectins A-chains (ML I-A, ML II-A, ML III-A and rML-A) have been obtained. In accordance to specificity of monAb they have been classified into three groups: 1. monAb against MLI-A and MLII-A, 2. monAb against MLII-A and MLIII-A, 3. monAb against A-subunits of MLI, MLII and MLIII. The results indicate, that antigen determinants of mistletoe lectins recognized by monAb MNA4, MNA9 and MTC12 do not contain any carbohydrates. Assay of lectins in mistletoe preparations was based on enzyme-linked lectin assay to meet the requirements given in the guidelines for drug tests. In this paper a sandwich ELISA test-system is described which allows to identify each of three ML-toxins. With the detection limits below 3 ng/ml and a linear measuring range of 3-30 ng/ml, dosages of mistletoe lectins in therapeutic range can be assayed. The problems in mistletoe lectins determination, structural differences and nature of heterogeneity of this proteins are discussed.