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Distribution of surface-exposed and non-accessible amino acid sequences among the two major structural domains of the S-layer protein of Aeromonas salmonicida.

Abstract：

:The tetragonally arranged crystalline surface protein array (A-layer) of the fish pathogenic bacterium Aeromonas salmonicida is a virulence factor. Circular dichroism studies in the presence or absence of 0.1% sodium dodecyl sulfate showed that the secondary structure of A-protein, and its 39,439 molecular weight amino-terminal trypsin-resistant peptide, were altered. In both cases alpha-helix was increased significantly at the expense of beta-structure when SDS was added. Western and dot immunoblotting, immuno-microscopy and enzyme-linked immunosorbent assay with monospecific polyclonal antiserum and eight monoclonal antibodies specific for epitopes exposed on the surface of native A-layer showed that the 481 residue A-protein subunit and the surface of A-layer were conserved antigenically. Mimeotope analysis of nonapeptides representing the sequence of A-protein allowed identification of 146 residues in presumed linear epitopes accessible on the surface of A-layer. Inaccessible or non-epitopic residues accounted for 70% of the protein. The majority of inaccessible residues were in the N-terminal 301 residues of A-protein. Dispersed among these were 65 surface-accessible residues in five linear epitope clusters illustrating the complex folding of this major structural domain of A-protein. The C-terminal 180 residues carried fewer linear epitopes but contained the major region of A-layers surface-accessible sequence, including four linear epitopes in predominantly hydrophobic sequence. Four A-layer surface-binding monoclonal antibodies also bound to this minor structural domain, although the epitopes of only two were identified by mimeotope analysis. The epitopes of six A-layer surface-binding monoclonals could not be identified, suggesting that A-layer may also contain conformation dependent surface epitopes.

journal_name

J Mol Biol

journal_title

Journal of molecular biology

authors

Doig P,McCubbin WD,Kay CM,Trust TJ

doi

10.1006/jmbi.1993.1550

subject

Has Abstract

pub_date

1993-10-20 00:00:00

pages

753-65

issue

eissn

0022-2836

issn

1089-8638

pii

S0022-2836(83)71550-0

journal_volume

233

pub_type

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Distribution of surface-exposed and non-accessible amino acid sequences among the two major structural domains of the S-layer protein of Aeromonas salmonicida.