BIOCHEMISTRY OF STREPTOCOCCUS MUTANS RIBOSOMES

  • Lee, John C (PI)

Project: Research project

Project Details

Description

Streptococcus mutans has been implicated as a major causative agent of
dental caries. Interference with this organism should markedly reduce
human caries. Ribosomal vaccines against S. mutans have been used
previously by other investigators but these ribosomal preparations usually
contain cell wall- and membrane-associated antigens and other antigens of
undefined nature which may cross-react with human tissues. It appears that
one of the major questions to be answered is that whether ribosomes and not
the contaminants are useful vaccines in the protection against dental
caries. If ribosomes are indeed useful vaccines, its mechanism of action
should be unique and warrents further investigation. Accordingly, the
specific aims of the present proposal are to prepare robosomes from S.
mutans of the highest degree of purity to exclude nonribosomal
contaminants. The purity of the ribosome will be monitored by biochemical
assays to assess the amount of protein, RNA, DNA and carbohydrate. The
sedimentation values of these purified ribosomes and their subnits will be
determined by sucrose density gradient centrifugation. The protein and RNA
composition of these highly purified ribosomes will be examined by
polycrylamide gel electrophoresis. Such highly purified ribosomes will be
used as immunogens to raise antibodies in rabbits. The specificity of
these antibodies will be tested against homologous S. mutans ribosomes and
whole cells as well as components of other serotypes using Ouchterlony
diffusion procedure and the Western blotting procedure. In addition,
specific RNA-protein complexes will be construced from purified ribosomal
RNA and proteins. Such complexes will be used as antigens. The antibodies
will also be tested for their effectiveness against S. mutans. The
rationale for using reconstituted ribonucleoprotein complexes is that the
chemical nature of the immunogen is defined and should not contain any
non-ribosomal contaminants. The specific RNA-protein complexes will
consist of 5S RNA and proteins from the purified large ribosomal subunit of
S. mutans. The identity of the proteins in the complex will also be
determined by 2-D polyacrylamide gel electrophoresis. If time permits
during this funding period, these highly purified ribosomes or RNA-protein
complexes will be administrated to animals and the salivary and serum
immune responses will be studied in order to test the potentials of using
these defined antigens as caries vaccines.
StatusFinished
Effective start/end date3/1/858/31/86

Funding

  • National Institutes of Health

ASJC

  • Medicine(all)
  • Dentistry(all)

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