Molecular cloning and expression of internalin in Listeria

International audienceThis chapter discusses the molecular cloning and expression of internalin in Listeria. Listeria monocytogenes is a ubiquitous gram-positive bacterium that causes severe infections in humans and in a variety of warm-blooded animals. This pathogen has emerged as an important agent of foodborne disease in the last decade. The majority of human infection affects pregnant women and result in stillbirths or neonatal sepsis. Meningitis, meningoencephalitis, and bacteremia account for most cases in nonpregnant adults occurring in the setting of immunosuppression or aging. The internalin (inl) region contains three open reading frames (ORFs): ORFA, inlA, and inlB. ORFA is 744 bp long and encodes a polypeptide of 248 amino acids that has 30% identity with the ribitol dehydrogenase from Klebsiella aerogenes, the protein encoded by the nodG gene of Rhizobium meliloti, and the actIII gene from Streptomyces coelicolor. inlA starts at 681 bp downstream from the end of ORFA, which is 2400 bp long. Two-thirds of inlA is made up of intragenic repeats that form two regions, region A and region B. These regions are independently formed by internal tandem duplications of ancestral sequences inlB starts 85 bp after the end of inlA and the two are separated by a putative transcription terminator, inlB is 1890 bp long and resembles inlA