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Linker Mutagenesis of the Caulobacter crescentus S-Layer Protein

  • Wade H. Bingle
  • Peter Awram
  • John Smit
Chapter
Part of the NATO ASI Series book series (NSSA, volume 252)

Abstract

As a part of its life cycle, the gram-negative bacterium Caulobacter crescentus exhibits a characteristic morphological switch between a sessile stalked cell and a flagellated dispersive or swarmer cell. During this complex differentiation process however, both cell types continue to elaborate a paracrystalline S-layer of hexagonal organization. The S-layer is composed of a single 98 kDa secreted protein (RsaA) noncovalently attached to other protein monomers and to the surface of the outer membrane; the latter interactions may be mediated by calcium ions and a specific Slayer associated molecule present in the outer membrane. Because RsaA is a secreted protein which interacts with itself as well as other molecules present in the outer membrane, multiple functional regions are expected to exist within the protein including those involved in secretion, calcium binding, outer membrane attachment and formation of the core and connectivity regions of the S-layer. Despite this expectation, analysis of the translated nucleotide sequence of the rsaA gene has not revealed possible functional regions of the protein beyond the existence of a probable calcium binding region. Similarly, N-terminal amino acid sequencing and sequencing of C-terminal peptide fragments derived from the S-layer protein have shown that no N- or C-terminal processing of the protein occurs, providing few clues to the mechanism of secretion. However, gene fusion studies have shown that the first 35–52 amino acids of the RsaA N-terminus can direct reporter proteins to the periplasm. (For a review of the present state of knowledge surrounding the C. crescentus S-layer, see Bingle et al., this book).

Keywords

Outer Membrane Inclusion Body Formation Swarmer Cell Caulobacter Crescentus Cover Amino Acid 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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References

  1. Bingle, W.H., and Smit, J., 1990, High-level expression vectors for Caulobacter crescentus incorporating the transcription/translation initiation regions of the paracrystalline surface-layer-protein gene, Plasmid 24: 143.PubMedCrossRefGoogle Scholar
  2. Bingle, W.H., and Smit, J., 1991, A method of tagging specific-purpose linkers with an antibiotic-resistance gene for linker mutagenesis using a selectable marker, Biotechniques 10: 150.PubMedGoogle Scholar
  3. Gilchrist, A., and Smit, J., 1992, Nucleotide sequence analysis of the gene encoding the Caulobacter crescentus paracrystalline surface layer, Can. J. Microbiol. 38: 193.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 1993

Authors and Affiliations

  • Wade H. Bingle
    • 1
  • Peter Awram
    • 1
  • John Smit
    • 1
  1. 1.Department of MicrobiologyUniversity of British ColumbiaVancouverCanada

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