Molecular analysis of Neisseria meningitidis class 3 outer membrane protein in strains recognized by the monoclonal antibody CB-Nm.2

Biotecnologia Aplicada 1997 Volume 14 No. 1, pp.23-30 Molecular analysis of Neisseria meningitidis class 3 outer membrane protein in strains recognized by the monoclonal antibody CB-Nm.2 Gerardo Guillen, Anabel lvarez, Consuelo Nazabal, Maria de Jesus Leal, Luis Manuel Alonso, Alexis Musacchio, Maite Delgado, Osvaldo Reyes and Ricardo Silva Code Number:BA97002 Sizes of files: Text: 48.8K Graphics: Line drawings (gif) -93K Photographs (jpg) - 22.8K ABSTRACT Bactericidal antibodies against outer membrane proteins are crucial to protect against Meningococcal Meningitis. The bactericidal monoclonal antibody (MAb) CB-Nm.2, specific for the class 3 outer membrane protein of Neisseria meningitidis, was assayed in an enzyme linked immunosorbent assay (ELISA) with a panel of 86 N. meningitidis strains. Fifty six strains belonging to seven serogroups: A, B, 29E, L, X, Y, Z, and five serotypes: 1, 4, 5, 12, 13, reacted with CB-Nm.2. The porB genes coding four such proteins were cloned and sequenced, and the translated amino acid sequences were compared with five previously published sequences. Sequence alignment revealed a five amino acid region (S/T)VETG located in the main variable region (VR) VR1 which was present in all N. meningitidis strains recognized by CB-Nm.2 and not in the strains which were negative in ELISA. Two synthetic peptides were designed on the basis of the predicted antigenic determinant for strains B385 and H355. Mouse antiserum obtained against the synthetic peptides recognized Neisseria strains in whole cell Dot-blot, but synthetic peptides failed to react with the MAb. The results show that the (S/T)VETG region is present among different serotypes of N. meningitidis and it is probably involved in antigenic recognition by the bactericidal MAb CB-Nm.2. Key words: class 3 protein, porB gene, DNA sequence RESUMEN Los anticuerpos bactericidas contra las proteinas de la membrana externa son decisivos en la proteccion contra la meningitis meningococcica. El anticuerpo monoclonal (AcM) CB-Nm.2 que es bactericida contra Neisseria meningitidis y especifico para la proteina de clase 3 de esta bacteria, se evaluo en ELISA contra un panel de 86 cepas de N. meningitidis. Cincuenta y seis de las cepas, pertenecientes a siete serogrupos: A, B, 29E, L, X, Y, Z, y cinco serotipos: 1, 4, 5, 12, 13, fueron reconocidas por el CB-Nm.2. Los genes porB codificantes para cuatro de estas proteinas se clonaron y secuenciaron. Las secuencias de ADN fueron traducidas a aminoacidos y comparadas con cinco secuencias publicadas con anterioridad. El alineamiento de secuencias mostro una region de cinco aminoacidos (S/T)VETG, localizada en la principal region variable (VR) VR1 que se encontraba presente en todas las cepas de N. meningitidis reconocidas por CB-Nm.2, y no en las cepas que resultaron negativas en el ELISA. Se sintetizaron peptidos que contenian esta region de las cepas B385 y H355. Los sueros de raton obtenidos contra los peptidos sinteticos reconocieron las cepas de Neisseria en Dot de celulas totales, pero los peptidos no fueron reconocidos por el AcM. Los resultados obtenidos muestran que la region (S/T)VETG es el posible sitio de reconocimiento antigenico del AcM bactericida CB-Nm.2 y esta presente en diferentes serotipos de N. meningitidis. Palabras claves: proteina de clase 3, gen porB, secuencia de ADN Copyright 1997 Elfos Scientia