RUI:  Ribosomal RNA Mutations and Ribosome Function
{'Code': '08070400', 'Directorate': {'Abbreviation': 'BIO', 'LongName': 'Direct For Biological Sciences'}, 'Division': {'Abbreviation': 'MCB', 'LongName': 'Div Of Molecular and Cellular Bioscience'}}
{'SignBlockName': 'Marcia Steinberg', 'PO_EMAI': None, 'PO_PHON': None}
The object of this project is:  to understand how the                           ribosome participates in the reactions of protein synthesis.                    The ribosome is a large complex ribonucleoprotein particle.                     The rRNA plays an important role in its function.  This                         includes base-pairing with mRNAs during translational                           initiation in prokaryotic systems, and possibly, other                          'catalytic' roles in the reactions of protein synthesis.                        These proposed interactions may involve a small region of                       primary structure or regions of RNA far apart on the primary                    structure, brought physically close by the higher order                         structure of the ribosome.  To investigate the relationship                     between the primary structure of rRNA and various functional                    roles, binding or catalytic, we are utilizing the procedures                    of site-specific mutagenesis.  Base changes are made to test                    if (a) the mutant rRNA will be incorporated into ribosomes                      and, if incorporated, (b) will they affect a specific                           reaction of protein synthesis.  Ideally a single base change                    in an important functional region will interfere with a                         unique stage of protein synthesis.  Ultimately, the                             identification of all bases involved in a unique function                       will establish the "active" site for that reaction.  Specific                   aims:  1. Production of site-specific mutations in 16S and                      23S rRNA at putative functional sites using a plasmid coded                     rrnB operron.  2. Synthesis of mutant ribosomes.  Mutant rDNA                   molecules in different plasmids will be transformed into E.                     coli, where specific in vivo functional tests will be carried                   out.  Mutant rRNA molecules will also be studied when under                     control of a number of non-rRNA promoters which allows the                      controlled expression of mutant rRNA molecules and allows for                   the study of mutations which when uncontrolled, are lethal to                   the cell.  3. Structural and functional assay of mutant                         ribosomes.  A. In vivo tests will include: (i) Tests for the                    effects on translation.  For example the mutant rRNA is                         introducted into an E. coli strain carrying a nonsense                          mutation in the Lac Z gene and a suppressor t-RNA.                              Alteration in the level of enzyme will be a measure of the                      effect of the mutant rRNA on translation.  (ii)  Association-                   Dissociation reaction: using the "maxicell" system we will                      determine if mutant 30S ribosomes will form 70S ribosomes,                      form polysomes, and recycle as cell-free extracts of E. coli                    and testing for their ability to carry out defined reactions                    of parotein synthesis.
{'FirstName': 'Melvin', 'LastName': 'Santer', 'PI_MID_INIT': None, 'PI_SUFX_NAME': None, 'PI_FULL_NAME': 'Melvin Santer', 'EmailAddress': None, 'NSF_ID': '000122362', 'StartDate': '04/01/1988', 'EndDate': None, 'RoleCode': 'Principal Investigator'}
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[{'Code': '1144', 'Text': 'Molecular Biophysics'}, {'Code': '9120', 'Text': 'CROSS-DIRECTORATE PROGRAMS'}]
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