Vam2/Vps41p and Vam6/Vps39p are components of a protein complex on the vacuolar membranes and involved in the vacuolar assembly in the yeast Saccharomyces cerevisiae

J Biol Chem. 1997 Apr 25;272(17):11344-9. doi: 10.1074/jbc.272.17.11344.

Abstract

The VAM2/VPS41 and VAM6/VPS39 were shown to encode hydrophilic proteins of 113 and 123 kDa, respectively. Deletion of the VAM2 and VAM6 functions resulted in accumulation of numerous vacuole-related structures of 200-400 nm in diameter that were much smaller than the normal vacuoles. Loss of functions of Vam2p and Vam6p resulted in inefficient processings of a set of vacuolar proteins, including proteinase A, proteinase B, and carboxypeptidase Y (CPY), and in severely defective maturation of another vacuolar protein, alkaline phosphatase. A part of newly synthesized CPY was missorted to the cell surface in the mutants. Epitope-tagged versions of Vam2p and Vam6p retained their functions, and they were found mostly in sedimentable fractions. The epitope-tagged Vam2p and Vam6p remained in the sedimentable fractions in the presence of Triton X-100, but they were extracted by urea or NaCl. Vam2p and Vam6p were cross-linked by the treatment of a chemical cross-linker. These observations indicated that Vam2p and Vam6p physically interact with each other and exist as components of a large protein complex. Vam6p fused with a green fluorescent protein were highly accumulated in a few specific regions of the vacuolar membranes. Large portions of Vam2p and Vam6p were fractionated into a vacuolar enriched fraction, indicating that they were localized mainly in the vacuolar membranes. These results showed that Vam2p and Vam6p execute their function in the vacuolar assembly as the components of a protein complex reside on the vacuolar membranes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Biological Transport
  • Carboxypeptidases / metabolism
  • Carrier Proteins / genetics*
  • Carrier Proteins / isolation & purification
  • Cathepsin A
  • Cell Compartmentation
  • Fungal Proteins / genetics*
  • Fungal Proteins / isolation & purification
  • Genes, Fungal
  • Genomic Library
  • Intracellular Membranes / chemistry
  • Intracellular Membranes / physiology*
  • Intracellular Membranes / ultrastructure
  • Membrane Proteins / genetics*
  • Membrane Proteins / isolation & purification
  • Molecular Sequence Data
  • Mutation
  • Nuclear Proteins*
  • Protein Precursors / metabolism
  • Protein Processing, Post-Translational / genetics
  • RNA-Binding Proteins / genetics*
  • RNA-Binding Proteins / isolation & purification
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae / ultrastructure
  • Saccharomyces cerevisiae Proteins*
  • Sequence Analysis, DNA
  • Vacuoles / chemistry
  • Vacuoles / genetics*
  • Vacuoles / ultrastructure
  • Vesicular Transport Proteins*

Substances

  • Carrier Proteins
  • Fungal Proteins
  • Membrane Proteins
  • NPL3 protein, S cerevisiae
  • Nuclear Proteins
  • Protein Precursors
  • RNA-Binding Proteins
  • Saccharomyces cerevisiae Proteins
  • VPS41 protein, S cerevisiae
  • Vesicular Transport Proteins
  • Carboxypeptidases
  • Cathepsin A

Associated data

  • GENBANK/AB000223
  • GENBANK/D83058