Effect of change in glycosylation of human serum transferrin on iron release to competing chelator EDTA

U ovom radu proučavan je utjecaj promjene glikozilacije transferina na konstantu vezanja željeza. Svrha rada bila je dokazati utjecaj glikanskog dijela strukture transferina na njegovu funkciju, točnije, na jačinu vezanja željeza. Sva ispitivanja su se provodila u PIPES puferu pri pH 7.4, na temperaturi 25°C, a kompleksirajući kompetitivni agens bila je EDTA. Korištena tehnika bila je spektroskopija u UV-Vis području. Iz podataka o apsorbanciji na 450 nm i koncentraciji dodane EDTA, izračunate su najprije pripadajuće koncentracije apo-transferina i holo-transferina te koncentracija otpuštenog željeza, a zatim i odgovarajuće konstante ravnoteže. Dobivene konstante ravnoteže veće su za red veličine kod vezanja željeza za nativni transferin u odnosu na asijalo-transferin. Također, kinetika otpuštanja željeza s asijalo-transferina je promijenjena u odnosu na nativnu sijaliniziranu formu.This work investigates the effect of change of transferrin glycosylation on the iron binding constant. The purpose of this study was to demonstrate the influence of the glycan part of the transferrin structure on its function, specifically on the iron binding strength. All experiments were carried out by UV-Vis spectroscopy in PIPES buffer (pH = 7.4) at 25° C and EDTA was used as a competitive chelator. Based on the absorbance data at 450 nm and the concentration of the added EDTA, concentrations of apo-transferin, holo-transferin and free iron were first obtained and then the corresponding equilibrium constants were calculated. The obtained equilibrium constants are an order of magnitude higher for the iron binding to the native transferrin, compared to asialo-transferrin. Additionally, the iron release kinetics from asialo-transferrin has been altered relative to the native sialylated protein form

Anti-TNF Biologicals Enhance the Anti-Inflammatory Properties of IgG N-Glycome in Crohn's Disease

Crohn’s disease (CD) is a chronic inflammation of the digestive tract that significantly impairs patients’ quality of life and well-being. Anti-TNF biologicals revolutionised the treatment of CD,yet many patients do not adequately respond to such therapy. Previous studies have demonstrated apro-inflammatory pattern in the composition of CD patients’ immunoglobulin G (IgG) N-glycomecompared to healthy individuals. Here, we utilised the high-throughput UHPLC method for N-glycan analysis to explore the longitudinal effect of the anti-TNF drugs infliximab and adalimumabon N-glycome composition of total serum IgG in 198 patients, as well as the predictive potential ofIgG N-glycans at baseline to detect primary non-responders to anti-TNF therapy in 1315 patients. Wediscovered a significant decrease in IgG agalactosylation and an increase in monogalactosylation,digalactosylation and sialylation during the 14 weeks of anti-TNF treatment, regardless of therapyresponse, all of which suggested a diminished inflammatory environment in CD patients treated withanti-TNF therapy. Furthermore, we observed that IgG N-glycome might contain certain informationregarding the anti-TNF therapy outcome before initiating the treatment. However, it is impossible to predict future primary non-responders to anti-TNF therapy based solely on IgG N-glycomecomposition at baseline

Susceptibility of Human Plasma N-glycome to Low-Calorie and Different Weight-Maintenance Diets

Aberrant plasma protein glycosylation is associated with a wide range of diseases, including diabetes, cardiovascular, and immunological disorders. To investigate plasma protein glycosylation alterations due to weight loss and successive weight-maintenance diets, 1850 glycomes from participants of the Diogenes study were analyzed using Ultra-High-Performance Liquid Chromatography (UHPLC). The Diogenes study is a large dietary intervention study in which participants were subjected to a low-calorie diet (LCD) followed by one of five different weight-maintenance diets in a period of 6 months. The most notable alterations of the plasma glycome were 8 weeks after the subjects engaged in the LCD; a significant increase in low-branched glycan structures, accompanied by a decrease in high-branched glycan structures. After the LCD period, there was also a significant rise in N-glycan structures with antennary fucose. Interestingly, we did not observe significant changes between different diets, and almost all effects we observed immediately after the LCD period were annulled during the weight-maintenance diets period

Susceptibility of Human Plasma N-glycome to Low-Calorie and Different Weight-Maintenance Diets

Aberrant plasma protein glycosylation is associated with a wide range of diseases, including diabetes, cardiovascular, and immunological disorders. To investigate plasma protein glycosylation alterations due to weight loss and successive weight-maintenance diets, 1850 glycomes from participants of the Diogenes study were analyzed using Ultra-High-Performance Liquid Chromatography (UHPLC). The Diogenes study is a large dietary intervention study in which participants were subjected to a low-calorie diet (LCD) followed by one of five different weight-maintenance diets in a period of 6 months. The most notable alterations of the plasma glycome were 8 weeks after the subjects engaged in the LCD; a significant increase in low-branched glycan structures, accompanied by a decrease in high-branched glycan structures. After the LCD period, there was also a significant rise in N-glycan structures with antennary fucose. Interestingly, we did not observe significant changes between different diets, and almost all effects we observed immediately after the LCD period were annulled during the weight-maintenance diets period

Effects of low-calorie and different weight-maintenance diets on IgG glycome composition

Obesity-induced inflammation activates the adaptive immune system by altering immunoglobulin G (IgG) glycosylation in a way to produce more proinflammatory antibodies. The IgG glycome has already been well studied, and its alterations are correlated with a high body mass index (BMI) and central adiposity. Still, the IgG N-glycome susceptibility to different dietary regimes for weight control after the initial weight loss has not been studied. To explore changes in IgG glycosylation induced by weight loss and subsequent weight-maintenance diets, we analyzed 1,850 IgG glycomes from subjects in a dietary intervention Diogenes study. In this study, participants followed a low-calorie diet (LCD) providing 800 kcal/d for 8 weeks, followed by one of five weight-maintenance diets over a 6-month period. The most significant alteration of the IgG N-glycome was present 8 weeks after the subjects underwent an LCD, a statistically significant decrease of agalactosylated and the increase of sialylated N glycans. In the follow-up period, the increase in glycans with bisecting GlcNAc and the decrease in sialylated glycans were observed. Those changes were present regardless of the diet type, and we did not observe significant changes between different diets. However, it should be noted that in all five diet groups, there were individuals who prominently altered their IgG glycome composition in either proinflammatory or anti-inflammatory directions

Immunoglobulin G glycome composition in transition from premenopause to postmenopause

Gonadal hormones affect immunoglobulin G (IgG) glycosylation, and the more proinflammatory IgG glycome composition might be one of the molecular mechanisms behind the increased proinflammatory phenotype in perimenopause. Using ultra-high-performance liquid chromatography, we analyzed IgG glycome composition in 5,080 samples from 1940 pre-, peri-, and postmenopausal women. Statistically significant decrease in galactosylation and sialylation was observed in postmenopausal women. Furthermore, during the transition from pre- to postmenopausal period, the rate of increase in agalactosylated structures (0.051/yr; 95%CI = 0.043–0.059, p < 0.001) and decrease in digalactosylated (−0.043/yr; 95%CI = −0.050 to −0.037, p < 0.001) and monosialylated glycans (−0.029/yr; 95%CI = −0.034 to −0.024, p < 0.001) were significantly higher than in either pre- or postmenopausal periods. The conversion to the more proinflammatory IgG glycome and the resulting decrease in the ability of IgG to suppress low-grade chronic inflammation may be an important molecular mechanism mediating the increased health risk in perimenopause and postmenopause

DataSheet_1_Effects of low-calorie and different weight-maintenance diets on IgG glycome composition.docx

Obesity-induced inflammation activates the adaptive immune system by altering immunoglobulin G (IgG) glycosylation in a way to produce more proinflammatory antibodies. The IgG glycome has already been well studied, and its alterations are correlated with a high body mass index (BMI) and central adiposity. Still, the IgG N-glycome susceptibility to different dietary regimes for weight control after the initial weight loss has not been studied. To explore changes in IgG glycosylation induced by weight loss and subsequent weight-maintenance diets, we analyzed 1,850 IgG glycomes from subjects in a dietary intervention Diogenes study. In this study, participants followed a low-calorie diet (LCD) providing 800 kcal/d for 8 weeks, followed by one of five weight-maintenance diets over a 6-month period. The most significant alteration of the IgG N-glycome was present 8 weeks after the subjects underwent an LCD, a statistically significant decrease of agalactosylated and the increase of sialylated N glycans. In the follow-up period, the increase in glycans with bisecting GlcNAc and the decrease in sialylated glycans were observed. Those changes were present regardless of the diet type, and we did not observe significant changes between different diets. However, it should be noted that in all five diet groups, there were individuals who prominently altered their IgG glycome composition in either proinflammatory or anti-inflammatory directions.</p

N-glycosylation of immunoglobulin G predicts incident hypertension

OBJECTIVES: Glycosylation of immunoglobulin G (IgG) is an important regulator of the immune system and has been implicated in prevalent hypertension.The aim of this study is to investigate whether the IgG glycome begins to change prior to hypertension diagnosis by analysing the IgG glycome composition in a large population-based female cohort with two independent replication samples. METHODS: We included 989 unrelated cases with incident hypertension and 1628 controls from the TwinsUK cohort (mean follow-up time of 6.3 years) with IgG measured at baseline by ultra-performance liquid chromatography and longitudinal BP measurement available. We replicated our findings in 106 individuals from the 10 001 Dalmatians and 729 from KORA S4. Cox regression mixed models were applied to identify changes in glycan traits preincident hypertension, after adjusting for age, mean arterial pressure, BMI, family relatedness and multiple testing (FDR &lt; 0.1). Significant IgG-incident hypertension associations were replicated in the two independent cohorts by leveraging Cox regression mixed models in the 10 001 Dalmatians and logistic regression models in the KORA cohort. RESULTS: We identified and replicated four glycan traits, incidence of bisecting GlcNAc, GP4, GP9 and GP21, that are predictive of incident hypertension after adjusting for confoundes and multiple testing [hazard ratio (95% CI) ranging from 0.45 (0.24-0.84) for GP21 to 2.9 (1.5-5.68) for GP4]. We then linearly combined the four replicated glycans and found that the glycan score correlated with incident hypertension, SBP and DBP. CONCLUSION: Our results suggest that the IgG glycome changes prior to the development of hypertension

Plasma protein N-glycome composition associates with postprandial lipaemic response

Abstract Background A dysregulated postprandial metabolic response is a risk factor for chronic diseases, including type 2 diabetes mellitus (T2DM). The plasma protein N-glycome is implicated in both lipid metabolism and T2DM risk. Hence, we first investigate the relationship between the N-glycome and postprandial metabolism and then explore the mediatory role of the plasma N-glycome in the relationship between postprandial lipaemia and T2DM. Methods We included 995 individuals from the ZOE-PREDICT 1 study with plasma N-glycans measured by ultra-performance liquid chromatography at fasting and triglyceride, insulin, and glucose levels measured at fasting and following a mixed-meal challenge. Linear mixed models were used to investigate the associations between plasma protein N-glycosylation and metabolic response (fasting, postprandial (C max), or change from fasting). A mediation analysis was used to further explore the relationship of the N-glycome in the prediabetes (HbA1c = 39–47 mmol/mol (5.7–6.5%))–postprandial lipaemia association. Results We identified 36 out of 55 glycans significantly associated with postprandial triglycerides (C max β ranging from -0.28 for low-branched glycans to 0.30 for GP26) after adjusting for covariates and multiple testing (p adjusted < 0.05). N-glycome composition explained 12.6% of the variance in postprandial triglycerides not already explained by traditional risk factors. Twenty-seven glycans were also associated with postprandial glucose and 12 with postprandial insulin. Additionally, 3 of the postprandial triglyceride–associated glycans (GP9, GP11, and GP32) also correlate with prediabetes and partially mediate the relationship between prediabetes and postprandial triglycerides. Conclusions This study provides a comprehensive overview of the interconnections between plasma protein N-glycosylation and postprandial responses, demonstrating the incremental predictive benefit of N-glycans. We also suggest a considerable proportion of the effect of prediabetes on postprandial triglycerides is mediated by some plasma N-glycans