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Biochemistry

Title:		Mutational analysis of the role of N-glycosylation in alpha-factor receptor function
Author(s):		Mentesana PE; Konopka JB;
Address:		"Department of Molecular Genetics and Microbiology, State University of New York, Stony Brook 11794-5222, USA"
Journal Title:		Biochemistry
Year:		2001
Volume:		40
Issue:		32
Page Number:		9685 - 9694
DOI:		10.1021/bi0108507
ISSN/ISBN:		0006-2960 (Print) 0006-2960 (Linking)
Abstract:		"The alpha-factor mating pheromone receptor (encoded by STE2) activates a G protein signaling pathway that stimulates the conjugation of Saccharomyces cerevisiae yeast cells. The alpha-factor receptor is known to undergo several forms of post-translational modification, including phosphorylation, mono-ubiquitination, and N-linked glycosylation. Since phosphorylation and mono-ubiquitination have been shown previously to play key roles in regulating the signaling activity and membrane trafficking of the alpha-factor receptors, the role of N-linked glycosylation was investigated in this study. The Asn residues in the five consensus sites for N-linked glycosylation present in the extracellular regions of the receptor protein were mutated to prevent carbohydrate attachment at these sites. Mutation of two sites near the receptor N-terminus (N25Q and N32Q) diminished the degree of receptor glycosylation, and the corresponding double mutant was not detectably N-glycosylated. The nonglycosylated receptors displayed normal function and subcellular localization, indicating that glycosylation is not important for wild-type receptor activity. However, mutation of the glycosylation sites resulted in improved plasma membrane localization for the Ste2-3 mutant receptors that are normally retained intracellularly at elevated temperatures. These results suggest that N-glycosylation may be involved in the sorting process for misfolded Ste2 proteins, and may similarly affect certain mutant receptors whose altered trafficking is implicated in human diseases"
Keywords:		"Amino Acid Sequence DNA Mutational Analysis Genes, Reporter Glycosylation Humans Immunoblotting Molecular Sequence Data Mutagenesis, Site-Directed Protein Structure, Secondary Receptors, Mating Factor Receptors, Peptide/chemistry/genetics/*metabolism Reco;"
Notes:		"MedlineMentesana, P E Konopka, J B eng R01 GM055107/GM/NIGMS NIH HHS/ 5T32CAO9176/CA/NCI NIH HHS/ GM55107/GM/NIGMS NIH HHS/ Research Support, U.S. Gov't, P.H.S. 2001/10/05 Biochemistry. 2001 Aug 14; 40(32):9685-94. doi: 10.1021/bi0108507"