Amino Acid Substitutions Affecting in Vivo Folding          Intermediates of the Thermostable P22 Tailspike Protein
{'Code': '08070400', 'Directorate': {'LongName': 'Direct For Biological Sciences'}, 'Division': {'LongName': 'Div Of Molecular and Cellular Bioscience'}}
{'SignBlockName': 'Eve Ida Barak'}
Some of the amino acid interactions critical for determining                    three dimensional protein structure must somehow direct the                     conformation of folding intermediates.  The thermostable                        tailspike endorhamnosidase of phage P22 has a thermolabile                      early intermediate in its folding and subunit association                       pathway.  Over 100 temperature sensitive folding mutations                      have been isolated which either further destabilize the                         thermolabile intermediate, or otherwise alter early kinetic                     steps.  Many of the substitutions of glycines and hydrophilic                   residues are at the surface of the native structure, probably                   associated with surface turns.  Since tailspike secondary                       structure is over 50% beta-sheet/turn, these are probably                       beta turns, and the mutations identify residues influencing                     their formation.                                                                                                                                                The specific objectives of the award are to:  1)  Identify                      the sites and amino acid substitutions for the remainder of                     the tsf mutations; 2)  For those substitutions altering                         charge, determine which are at the surface and therefore turn                   candidates, and analyze the folding defect in these mutants;                    3)  Isolate second site suppressors of these defects, to                        determine whether the interacting residues are local or                         distant in the chain; 4) Isolate and characterize the                           protrimer intermediate from wild type and mutant infected                       cells; 5)  Isolate and characterize the aggregated chains                       that accumulate at high temperature as a means of                               investigating the conformation of the early intermediate from                   which it forms; 6)  Purify tsf mutant and tsf/suppressor-tsf                    double mutant proteins for collaborative X-ray diffraction,                     Raman spectroscopy, in vitro refolding, and differential                        scanning calorimetry studies.
{'FirstName': 'Jonathan', 'LastName': 'King', 'EmailAddress': 'jaking@mit.edu', 'StartDate': '08/01/1987', 'EndDate': None, 'RoleCode': 'Principal Investigator'}
{'Name': 'Massachusetts Institute of Technology', 'CityName': 'Cambridge', 'ZipCode': '021394301', 'PhoneNumber': '6172531000', 'StreetAddress': '77 MASSACHUSETTS AVE', 'CountryName': 'United States', 'StateName': 'Massachusetts', 'StateCode': 'MA'}
{'Code': '1144', 'Text': 'Molecular Biophysics'}
{'url': 'https://www.nsf.gov/awardsearch/download?DownloadFileName=1987&All=true', 'xml': '8704126.xml'}