Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.5.1.52 (PNGase F)
 1,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Purified plasma membranes from the yeast Saccharomyces cerevisiae bind about 1.2 pmol of cAMP/mg of protein with high affinity (Kd = 6 nM). By using photoaffinity labeling with 8-N3-[32P]cAMP, we have identified in plasma membrane vesicles a cAMP-binding protein (Mr = 54,000) that is present also in bcy1 disruption mutants, lacking the cytoplasmic R subunit of protein kinase A (PKA). This argues that it is genetically unrelated to PKA. Neither high salt, nor alkaline carbonate, nor cAMP extract the protein from the membrane, suggesting that it is not peripherally bound. The observation that (glycosyl)phosphatidylinositol-specific phospholipases (or nitrous acid) release the amphiphilic protein from the membrane, thereby converting it to a hydrophilic form, indicates anchorage by a glycolipidic membrane anchor. Treatment with N-glycanase reduces the Mr to 44,000-46,000 indicative of a modification by N-linked carbohydrate side chain(s). In addition to the action of a phospholipase, the efficient release from the membrane requires the removal of the carbohydrate side chain(s) or the presence of high salt or methyl alpha-mannopyranoside, suggesting complex interactions with the membrane involving not only the glycolipidic anchor but also the glycan side chain(s). Topological studies show that the protein is exposed to the periplasmic space, raising intriguing questions for the function of this protein.
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PMID:A cAMP-binding ectoprotein in the yeast Saccharomyces cerevisiae. 165 42

Variable amounts of soluble forms of a variety of glycosyl-phosphatidylinositol (GPI)-anchored proteins occur extracellularly, but the molecular mechanisms governing their release are not entirely clear. When the GPI-anchored folate receptor (FR) type beta was expressed transiently in human 293 fibroblasts, there was a roughly equal distribution of [3H]folic acid binding protein between the cell surface and the medium after 24 h over a wide range of expression levels of FR-beta. The difference in apparent molecular masses between the soluble FR-beta and the PI-PLC-treated membrane protein indicated that the former was not released from the membrane by the action of phospholipase. Brefeldin A inhibited the release of soluble FR-beta from both the transfected 293 cells and stable recombinant CHO (CHO-FR-beta) cells while pre-existing levels of cell surface FR were unaltered suggesting the absence of a precursor-product relationship between the membrane-associated FR-beta and the soluble protein in the medium. [35S]Cysteine pulse-chase analysis was consistent with this finding. Interchanging of carboxyl-terminal peptides between FR-beta and FR-alpha revealed that the nature of the processed signal for GPI modification was responsible for the quantitative membrane anchoring of FR-alpha and the production of soluble FR-beta. When total cell lysates were analyzed by Western blot, a diffuse band of apparent 41 kDa and three additional sharp bands of apparent 35, 33, and 29.3 kDa were seen. The 41 kDa band was identified as the PI-PLC sensitive cell surface receptor. Several mutant constructs of FR-beta, in which the carboxyl-terminal signal for GPI modification was either disrupted or deleted only gave the three lower bands. The three sharp bands from the wild-type and the mutant forms of FR-beta were identified as nonglycosylated (29.3 kDa) or glycosylated polypeptides in which the carboxyl-terminal peptide was at least partially proteolyzed without GPI modification. All of the mutations in the GPI signal resulted in the recovery of [3H]folic acid binding protein in the media which, similar to the wild-type FR recovered from the media, were converted to the 29.3 kDa band by N-glycanase. The results from this study indicate that a carboxyl-terminal peptide in FR-beta is efficiently proteolyzed intracellularly by a pathway that is independent of GPI signal recognition resulting in proper protein folding and secretion. Such carboxyl-terminal sequences could represent a simple adaptation for proteins whose physiologic functions reside both at the cell surface and in extracellular fluids, allowing their selective and tissue-specific release.
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PMID:Proteolysis of the carboxyl-terminal GPI signal independent of GPI modification as a mechanism for selective protein secretion. 939 77