TI  - Identification of an 11-kDa <prot>FKBP12</prot>-rapamycin-binding domain within the
      289-kDa <prot>FKBP12-rapamycin-associated protein</prot> and characterization of a
      critical serine residue.
PG  - 4947-51
AB  - Complexed with its intracellular receptor,  <prot>FKBP12</prot> , the natural product
       rapamycin  inhibits G1 progression of the cell cycle in a variety of
      mammalian cell lines and in the yeast Saccharomyces cerevisae. Previously,
      a mammalian protein that directly associates with  <prot>FKBP12</prot> - rapamycin  has
      been identified and its encoding gene has been cloned from both human
      (designated  <prot>FRAP</prot> ) [Brown, E.J., Albers, M.W., Shin, T.B., Ichikawa, K.,
      Keith, C.T., Lane, W.S. &amp; Schreiber, S.L. (1994) Nature (London) 369,
      756-758] and rat (designated <prot>RAFT</prot>) [Sabatini, D.M., Erdjument-Bromage, H.,
      Lui, M., Tempst, P. &amp; Snyder, S.H. (1994) Cell 78, 35-43]. The full-length
      <prot>FRAP</prot> is a 289-kDa protein containing a putative phosphatidylinositol
      kinase domain. Using an in vitro transcription/translation assay method
      coupled with proteolysis studies, we have identified an 11-kDa
       <prot>FKBP12</prot>-rapamycin -binding domain within  <prot>FRAP</prot>. This minimal binding domain
      lies N-terminal to the kinase domain and spans residues 2025-2114. In
      addition, we have carried out mutagenesis studies to investigate the role
      of Ser2035, a potential phosphorylation site for <prot>protein kinase C</prot> within
      this domain. We now show that the <prot>FRAP</prot> Ser2035--&gt;Ala mutant displays
      similar binding affinity when compared with the wild-type protein, whereas
      all other mutations at this site, including mimics of phosphoserine,
      abolish binding, presumably due to either unfavorable steric interactions
      or induced conformational changes.
AD  - Howard Hughes Medical Institute, Department of Chemistry, Harvard
