INVESTIGADORES
D'ALESSIO Cecilia
congresos y reuniones científicas
Título:
Development of a molecular tool to measure in vivo protein hypoglycosylation in fission yeasts
Autor/es:
GALLO, GL; HERRERA AGUILAR, N; ORSI, R.; MATTERA, V; PARODI, AJ; D'ALESSIO, C
Lugar:
Rosario
Reunión:
Congreso; L reunión anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2014
Institución organizadora:
SAIB
Resumen:
¨
During N-glycosylation the endoplasmic
reticulum (ER) membrane oligosaccharytransferase complex (OST) transfers Glc3Man9GlcNAc2
from a dolichol-PP donor to the sequence Asn-X-Ser/Thr
of proteins that are entering into the ER. The addition of a bulky hydrophilic
glycan enhances protein folding efficiency as it reduces aggregation of folding
intermediates. Defects in the glycan transfer reaction due either to OST
mutations or to truncated glycan structures may result in protein
hypoglycosylation (lack of glycosylation of sites normally occupied) thus
causing ubiquitous defects and in human diseases known as congenital disorders
of glycosylation type I. It has been recently shown that a GFP variant in which
an N-glycosylation site was
introduced (Gly-GFP) loses fluorescence when such site is occupied by a glycan.
We expressed GFP and Gly-GFP variants fused to an N-terminal S. pombe signal peptide and a C-terminal
ER retention signal (ER-GFP and ER-GlyGFP), both in a wild type and in an Dalg6 mutant in which hypoglycosylation
occurs. Our results showed that while expressed ER-GFP fluoresces in the ER of
both strains, ER-GlyGFP only fluoresces in the ER of Dalg6 mutant, indicating that the fluorescence intensity of
this construction may be used to test the glycoprotein hypoglycosylation in vivo in S. pombe.