Assuntos
Proteínas de Bactérias/toxicidade , Escherichia coli/genética , Genes Bacterianos/genética , Plasmídeos/genética , Regiões Promotoras Genéticas/genética , Arabinose/farmacologia , Clonagem Molecular , Expressão Gênica/genética , Isopropiltiogalactosídeo/farmacologia , Lactose/farmacologia , Dados de Sequência Molecular , Proteínas Recombinantes/isolamento & purificaçãoRESUMO
Mature beta-lactamase was attached to the N-terminus of human glycophorin C, an N-out membrane protein lacking a cleavable signal peptide (an N-tail membrane protein). When synthesised in Escherichia coli more than 30% of the intact mature beta-lactamase-glycophorin C molecules assembled N-out, C-in into the cytoplasmic membrane. The N-tail translocated beta-lactamase folded into an enzymatically active form, but it was more susceptible to proteolysis than the equivalent portion of beta-lactamase-glycophorin C synthesised with an N-terminal signal peptide. Its translocation was virtually abolished when the N-out domain of glycophorin C was truncated or when the basic residues C-terminally flanking the glycophorin C membrane-spanning segment were replaced with neutral ones.
