IDENTIFICATION OF 3 MANNOPROTEINS IN THE CELL-WALL OF SACCHAROMYCES-CEREVISIAE

Three glucanase-extractable cell wall proteins from Saccharomyces cere visiae were purified, and their N-terminal amino acid sequences were d etermined. With this information, we were able to assign gene products to three known open reading frames (ORFs). The N-terminal sequence of a 55-kDa mannoprotein corresponded with the product of ORF YKL096w, w hich we named CWP1 (cell wall protein 1). A 80-kDa mannoprotein was id entified as the product of the TIP1 gene, and a 180-kDa mannoprotein c orresponded to the product of the ORF YKL444, which we named CWP2. CWP 1, TIP1, and CWP2 encode proteins of 239, 210, and 92 amino acids, res pectively. The C-terminal regions of these proteins all consist for mo re than 40% of serine/threonine and contain putative glycosylphosphati dylinositol attachment signals. Furthermore, Cwp1p and Tip1p were show n to carry a beta 1,6-glucose-containing side chain. The cwp2 deletion mutant displayed an increased sensitivity to Congo red, calcofluor wh ite, and Zymolyase. Electron microscopic analysis of the cwp2 deletion mutant showed a strongly reduced electron-dense layer on the outside of the cell wall. These results indicate that Cwp2p is a major constit uent of the cell wall and plays an important role in stabilizing the c ell wall. Depletion of Cwp1p or Tip1p also caused increased sensitivit ies to Congo red and calcofluor white, but the effects were less prono unced than for cwp2 Delta. All three cell wall proteins show a substan tial homology with Srp1p, which also appears to be localized in the ce ll wall. We conclude that these four proteins are small structurally r elated cell wall proteins.