The Compartmentalisation of Phosphorylated Free Oligosaccharides in Cells from a CDG Ig Patient Reveals a Novel ER-to-Cytosol Translocation Process

BACKGROUND: Biosynthesis of the dolichol linked oligosaccharide (DLO) required for protein N-glycosylation starts on the cytoplasmic face of the ER to give Man(5)GlcNAc(2)-PP-dolichol, which then flips into the ER for further glycosylation yielding mature DLO (Glc(3)Man(9)GlcNAc(2)-PP-dolichol). After transfer of Glc(3)Man(9)GlcNAc(2) onto protein, dolichol-PP is recycled to dolichol-P and reused for DLO biosynthesis. Because de novo dolichol synthesis is slow, dolichol recycling is rate limiting for protein glycosylation. Immature DLO intermediates may also be recycled by pyrophosphatase-mediated cleavage to yield dolichol-P and phosphorylated oligosaccharides (fOSGN2-P). Here, we examine fOSGN2-P generation in cells from patients with type I Congenital Disorders of Glycosylation (CDG I) in which defects in the dolichol cycle cause accumulation of immature DLO intermediates and protein hypoglycosylation. METHODS AND PRINCIPAL FINDINGS: In EBV-transformed lymphoblastoid cells from CDG I patients and normal subjects a correlation exists between the quantities of metabolically radiolabeled fOSGN2-P and truncated DLO intermediates only when these two classes of compounds possess 7 or less hexose residues. Larger fOSGN2-P were difficult to detect despite an abundance of more fully mannosylated and glucosylated DLO. When CDG Ig cells, which accumulate Man(7)GlcNAc(2)-PP-dolichol, are permeabilised so that vesicular transport and protein synthesis are abolished, the DLO pool required for Man(7)GlcNAc(2)-P generation could be depleted by adding exogenous glycosylation acceptor peptide. Under conditions where a glycotripeptide and neutral free oligosaccharides remain predominantly in the lumen of the ER, Man(7)GlcNAc(2)-P appears in the cytosol without detectable generation of ER luminal Man(7)GlcNAc(2)-P. CONCLUSIONS AND SIGNIFICANCE: The DLO pools required for N-glycosylation and fOSGN2-P generation are functionally linked and this substantiates the hypothesis that pyrophosphatase-mediated cleavage of DLO intermediates yields recyclable dolichol-P. The kinetics of cytosolic fOSGN2-P generation from a luminally-generated DLO intermediate demonstrate the presence of a previously undetected ER-to-cytosol translocation process for either fOSGN2-P or DLO

Synthetic strategies to nanostructured photocatalysts for CO2 reduction to solar fuels and chemicals

Artificial photosynthesis represents one of the great scientific challenges of the 21st century, offering the possibility of clean energy through water photolysis and renewable chemicals through CO2 utilisation as a sustainable feedstock. Catalysis will undoubtedly play a key role in delivering technologies able to meet these goals, mediating solar energy via excited generate charge carriers to selectively activate molecular bonds under ambient conditions. This review describes recent synthetic approaches adopted to engineer nanostructured photocatalytic materials for efficient light harnessing, charge separation and the photoreduction of CO2 to higher hydrocarbons such as methane, methanol and even olefins