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Molecular requirements for attachment of the glycosylphosphatidylinositol anchor to the human alpha folate receptor
Author(s) -
Tomassetti Antonella,
Bottero Federica,
Mazzi Mimma,
Miotti Silvia,
Colnaghi Maria I.,
Canevari Silvana
Publication year - 1999
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/(sici)1097-4644(19990101)72:1<111::aid-jcb12>3.0.co;2-1
Subject(s) - complementary dna , folate receptor , biochemistry , biology , amino acid , microbiology and biotechnology , gene isoform , peptide sequence , chinese hamster ovary cell , arginine , threonine , transfection , cleavage (geology) , c terminus , site directed mutagenesis , receptor , serine , phosphorylation , mutant , gene , paleontology , genetics , cancer , cancer cell , fracture (geology)
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 Ser 234 with arginine or threonine, or replacement of Gly 235 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 Ser 234 is not the only potential cleavage/attachment site of the α isoform of FR. J. Cell. Biochem. 72:111–118, 1999. © 1999 Wiley‐Liss, Inc.