Meningococcus adhesins nada, app and orf 40

Abstract

NadA, App and ORF40 function as adhesins in N. meningitidis. Adhesion can be modulated by targeting these three proteins. NadA allelic variants are disclosed. Autoproteolytic cleavage of App is disclosed, as is removal of the activity by mutagenesis. App is processed and secreted into culture medium when expressed in E. coli. Mature App proteins are disclosed. Knockout mutants are disclosed. Vesicles from non-Neisserial hosts with heterologous adhesin expression are disclosed.

Description

[0001] All documents cited herein are incorporated by reference in their entirety. TECHNICAL FIELD [0002] This invention is in the field of biochemistry and, in particular, the biochemistry of the pathogenic bacteria in the genus Neisseria (e.g. N. meningitidis and N. gonorrhoeae). BACKGROUND ART [0003] International patent applications WO99 / 24578, WO99 / 36544, WO99 / 57280 and WO00 / 22430 disclose proteins from Neisseria meningitidis and Neisseria gonorrhoeae. The complete genome sequence of serogroup B N. meningitidis has been published [Tettelin et al. (2000) Science 287:1809-1815) and has been subjected to analysis in order to identify vaccine antigens [Pizza et al. (2000) Science 287:1816-1820]. Approaches to expression of the proteins are disclosed in WO01 / 64922. The complete genome sequence of serogroup A N. meningitidis is also known (Parchill et al. (2000) Nature 404:502-506]. [0004] Sequence data alone, however, does not reveal everything about this pathogen. Objects of the pr...

AI Technical Summary

Benefits of technology

[0044] The invention provides a method for preventing the attachment of a Neisserial cell to an epithelial cell, wherein one or more of App, ORF40 and / or NadA is knocked out.

[0048] The knockout mutation will reduce the level of mRNA encoding App, ORF40 and / or NadA to <1% of that produced by the wild-type bacterium, preferably <0.5%, more preferably <0.1%, and most preferably to 0%.

[0071] International patent application WO01 / 52885 discloses that the addition of further defined components to OMV vaccines significantly broadens their efficacy.

[0073] It has now been found that OMVs prepared from E. coli which express a heterologous Neisseria gene can give better results in standard immunogenicity tests than the antigens in purified form.

[0113] Preferred fragments within category (b) lack the N-terminal leader peptide. For SEQ IDs 1, 2, 3, 7, 9, 11 & 13 the value of k is thus 23; for SEQ IDs 4, 5, 6, 8, 10, 12 & 14 the value of k is 25. The leader peptide may be replaced with the leader peptide from another protein, by another protein (i.e. to form a fusion protein) or by an alternative N-terminus sequence to allow efficient expression.

[0125] Mutants can include amino acid substitutions, additions or deletions. The amino acid substitutions can be conservative amino acid substitutions or substitutions to eliminate non-essential amino acids, such as to alter a glycosylation site, a phosphorylation site or an acetylation site, or to minimize misfolding by substitution or deletion of one or more cysteine residues that are not necessary for function. Conservative amino acid substitutions are those that preserve the general charge, hydrophobicity / hydrophilicity, and / or steric bulk of the amino acid substituted. Variants can be designed so as to retain or have enhanced biological activity of a particular region of the polypeptide (e.g. a functional domain and / or, where the polypeptide is a member of a polypeptide family, a region associated with a consensus sequence). Selection of amino acid alterations for production of variants can be based upon the accessibility (interior vs. exterior) of the amino acid, the thermostability of the variant polypeptide, desired disulfide bridges, desired metal binding sites etc.

Problems solved by technology

Sequence data alone, however, does not reveal everything about this pathogen.

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