Molecular requirements for attachment of the glycosylphosphatidylinositol anchor to the human alpha folate receptor

Antonella Tomassetti, Federica Bottero, Mimma Mazzi, Silvia Miotti, Maria I. Colnaghi, Silvana Canevari

Research output: Contribution to journalArticlepeer-review

Abstract

The α isoform of the folate receptor (FR) is a 38-KDa glycosylphosphatidylinositol (GPI) protein which mediates the internalization of folates. The FR amino acid sequence has features typical of GPI-linked proteins, including the presence of a hydrophobic carboxyl-terminus, a hinge region, and a stretch of small and uncharged amino acids. Substitution of predicted cleavage/attachment Ser234 with arginine or threonine, or replacement of Gly235 with proline by site-directed mutagenesis had no effect on GPI processing. In fact, CHO cells transfected with each of the three cDNA variants or with FR wild-type showed comparable amounts of phosphatidylinositol-specific phospholipase C-resistant FR in double- determinant radioimmunoassay. Western blot analysis of total cell lysates from all transfectants consistently revealed the 38-KDa FR band. Deletion of residues 233-237 in the amino-terminal portion of the FR cDNA constructs derived by a polymerase chain reaction strategy abrogated GPI processing, with only a small proportion of the FR remaining in the cytoplasm in four of the five clones tested. This finding suggests that FR residues 233-237 are essential in properly juxtaposing the FR hydrophobic domain. Together, these data support the hypothesis that the postulated Ser234 is not the only potential cleavage/attachment site of the α isoform of FR.

Original languageEnglish
Pages (from-to)111-118
Number of pages8
JournalJournal of Cellular Biochemistry
Volume72
Issue number1
DOIs
Publication statusPublished - Jan 1 1999

Keywords

  • Alpha-folate receptor
  • Cleavage/attachment site
  • Glycosylphosphatidylinositol anchor
  • Membrane receptor
  • Site-directed mutagenesis

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology

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