Functional analysis of Ost3p and Ost6p containing yeast oligosaccharyltransferases.

The oligosaccharyltransferase (OST) is the central enzyme in the N-glycosylation pathway. It transfers a defined oligosaccharide from a lipid-linker onto the asparagine side chain of proteins. The yeast OST consists of eight subunits and exists in two catalytically distinct isoforms that differ in one subunit, Ost3p or Ost6p. The cryo-electron microscopy structure of the Ost6p containing complex was found to be highly similar to the Ost3p containing OST. OST enzymes with altered Ost3p/Ost6p subunits were generated and functionally analyzed. The three C-terminal transmembrane helices were responsible for the higher turnover-rate of the Ost3p vs. the Ost6p containing enzyme in vitro and the more severe hypoglycosylation in Ost3p lacking strains in vivo. Glycosylation of specific OST target sites required the N-terminal thioredoxin domain of Ost3p or Ost6p. This Ost3p/Ost6p dependence was glycosylation site but not protein specific. We concluded that the Ost3p/Ost6p subunits modulate the catalytic activity of OST and provide additional specificity for OST substrate recognition.

Urinary Biomarkers in the Prediction of Prognosis and Treatment Response in IgA Nephropathy.

BACKGROUND/AIMS: The addition of immunosuppression to supportive care reduces proteinuria in a subset of patients with IgA nephropathy (IgAN) but is associated with an increased rate of adverse events. The present work investigates whether urinary biomarkers are able to identify subjects who benefit from immunosuppression and to predict the progression of disease in a sub-cohort of the STOP-IgAN trial. METHODS: Urinary neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), calprotectin, and the product of tissue inhibitor of metalloproteinase-2 and insulin-like growth factor-binding protein 7 (TIMP2•IGFBP7) were measured in all available urine samples obtained at the time point of enrollment in the STOP-IgAN trial (n=113). RESULTS: Biomarker concentrations in both the overall study population and the subgroup with additional immunosuppression did not differ in subjects reaching vs. not reaching full clinical remission, eGFR loss ≥ 15, or 30 ml/min/1.73 m2 over the 3-year trial phase (p> 0.05 each). Receiver-operating characteristic curves showed a poor predictive accuracy of each biomarker for the above-mentioned parameters in the overall study population (areas under the curve ≤0.611). Accordingly, there was neither a significant correlation of any biomarker and adverse outcome in linear regression analysis, nor between biomarker concentrations at enrollment and change in the eGFR over the 3-year observation period. CONCLUSION: NGAL, KIM-1, calprotectin, and [TIMP-2]•[IGFBP7] had neither a prognostic value for the progression of IgAN, nor for the response to immunosuppression in the present sub-cohort of the STOP-IgAN trial. The search for appropriate biomarkers for an individualized treatment strategy in IgAN continues.