Complementation analysis of Pasteurella haemolytica leukotoxin deletion proteins /
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| Other Authors: | , |
| Format: | Thesis Book |
| Language: | English |
| Published: |
1991.
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| Subjects: | |
| Online Access: | Link to OAKTrust copy |
| Abstract: | A series of in-frame deletions in the leukotoxin IktA gene of Pasteurella haemolytica was constructed. All of the deletions eliminated the lytic activity of the leukotoxin towards the bovine lymphoma cell line, BL3. Deletions which removed segments of the amino-terminal hydrophobic region, which has been regarded as the lytic domain because of its membrane-seeking properties, were found to agglutinate BL3 cells. Agglutination was similar to lysis by the wild-type toxin in that it was cell-type specific, and required the presence of calcium and IktC gene expression. The agglutinating deletion proteins protected BL3 cells from lysis by the wild-type toxin in a competitive fashion. This suggests that the agglutinating mutant proteins bind to a surface feature of the bovine leukocyte which interacts with the native leukotoxin. Thus, the cell-binding domain is located in the carboxy-terminal half of LktA and can function independently of the amino-terminal domain. The agglutinating deletion proteins were able to complement derivatives of LktA with deletions distal to the amino-terminal hydrophobic regions. This suggests that the lytic and cell-binding domains of LktA reside in the amino- and carboxy-terminal halves of the toxin, respectively, and are functionally independent. LktC-activation is required only for the optimal binding of the toxin to the target cells and not for the function of the pore-forming region of LktA. Immunoprecipitation experiments provided evidence that the reconstitution of lytic ability by complementation between inactive LktA deletion proteins is the result of hetero-oligomer formation, in which one protein supplies the functional leukocyte-binding domain and the second protein provides the lytic domain. On the basis of complementation and immunoprecipitation results, a putative oligomerization domain is assigned to the central region, flanking codon 548, of the LktA protein. |
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| Item Description: | Typescript (photocopy). Vita. "Major subject: Microbiology." |
| Physical Description: | xi, 137 leaves : illustrations ; 29 cm |
| Bibliography: | Includes bibliographical references. |