Ser787 in the proline-rich region of human MAP4 is a critical phosphorylation site that reduces its activity to promote tubulin polymerization

p34(cdc2) kinase-phosphorylation sires in the microtubule (MT)-binding regi on of MAP4 were determined by peptide sequence of phosphorylated MTB3, a fr agment containing the carboxy-terminal half of human MAP4. In addition to t wo phosphopeptides containing Ser696 and Ser787 which were previously indic ated to be in vivo phosphorylation sites, two novel phosphopeptides, contai ning Thr892 or Thr901 and Thr917 as possible phosphorylation sites, were is olated, though only in in vitro phosphorylation. The role of phosphorylatio n at Ser696 and Ser787, which were differently phosphorylated during the ce ll cycle (Ookata et al., (1997), Biochemistry: 36: 15873-15883), was invest igated in MT-polymerization, using MAP4 Ser to Glu mutants, which mimic pho sphorylation at each site. Mutation of Ser787 to Glu strikingly reduced the MAP4's MT-polymerization activity, while Glu-mutation at Ser696 did not. T hese results suggest that Ser787 could be the critical phosphorylation site causing MTs to be dynamic at mitosis.