Human fructosamine-3-kinase: purification, sequencing, substrate specificity, and evidence of activity in vivo.
ABSTRACT Nonenzymatic glycation appears to be an important factor in the pathogenesis of diabetic complications. Key early intermediates in this process are fructosamines, such as protein-bound fructoselysines. In this report, we describe the purification and characterization of a mammalian fructosamine-3-kinase (FN3K), which phosphorylates fructoselysine (FL) residues on glycated proteins, to FL-3-phosphate (FL3P). This phosphorylation destabilizes the FL adduct and leads to its spontaneous decomposition, thereby reversing the nonenzymatic glycation process at an early stage. FN3K was purified to homogeneity from human erythrocytes and sequenced by means of electrospray tandem mass spectrometry. The protein thus identified is a 35-kDa monomer that appears to be expressed in all mammalian tissues. It has no significant homology to other known proteins and appears to be encoded by genomic sequences located on human chromosomes 1 and 17. The lability of FL3P, the high affinity of FN3K for FL, and the wide distribution of FN3K suggest that the function of this enzyme is deglycation of nonenzymatically glycated proteins. Because the condensation of glucose and lysine residues is an ubiquitous and unavoidable process in homeothermic organisms, a deglycation system mediated by FN3K may be an important factor in protecting cells from the deleterious effects of nonenzymatic glycation. Our sequence data of FN3K are in excellent agreement with a recent report on this enzyme by Delpierre et al. (Diabetes 49:1627-1634, 2000).
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ABSTRACT: The non-enzymatic reaction between glucose and protein can be chemically reversed by transglycation. Here we report the transglycation activity of hydralazine using a newly developed MALDI-TOF-MS based assay. Hydralazine mediated transglycation of HbA1c, plasma proteins and kidney proteins was demonstrated in streptozotocin (STZ) induced diabetic mice, as evidenced by decrease in protein glycation, as well as presence of hydralazine-glucose conjugate in urine of diabetic mice treated with hydralazine. Hydralazine down regulated the expression of Receptor for Advanced Glycation End products (RAGE), NADPH oxidase (NOX), and super oxide dismutase (SOD). These findings will provide a new dimension for developing intervention strategies for the treatment of glycation associated diseases such as diabetes complications, atherosclerosis, and aging.Scientific Reports 01/2013; 3:2941. · 5.08 Impact Factor
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ABSTRACT: OBJECTIVE — To assess the relative importance of fasting and postprandial hyperglycemia to vascular dysfunction in diabetes, we have measured indicators of glycation, oxidative and nitrosative stress in subjects with type 1 diabetes, and different postprandial glucose patterns. RESEARCH DESIGN AND METHODS — Plasma and urinary levels of specific argi-nine-and lysine-derived advanced glycation end products, as well as oxidative and nitrosative products, were measured by liquid chromatography with triple quadrupole mass spectrometric detection (LC-MS/MS) after 2 months of treatment with insulin lispro or human regular insulin in 21 subjects participating in a cross-over study. Hb-bound early glycation (Amadori) products were also measured after each treatment period by high-performance liquid chromatography (fructosyl-valine Hb or HbA 1c [A1C]:Diamat) and fructosyl-lysine Hb by LC-MS/MS (A1C: fructosyl-lysine). RESULTS — In diabetic patients, the concentrations of protein glycation and oxidation-free adducts increased up to 10-fold, while urinary excretion increased up to 15-fold. Decreasing postprandial hyperglycemia with lispro gave 10 –20% decreases of the major free glycation adducts, hydroimidazolones derived from methylglyoxal and 3-deoxyglucosone, and glyoxal-derived Nε-carboxymethyl-lysine. No differences were observed in A1C:Diamat or A1C: fructosyl-lysine with lispro or regular insulin therapy in spite of significant decreases in postprandial glycemia with lispro. CONCLUSIONS — We conclude that the profound increases in proteolytic products of proteins modified by advanced glycation endproducts in diabetic patients are responsive to changes in mean hyperglycemia and also show responses to changes in postprandial hyperglycemia.
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ABSTRACT: In our previous publication we reported on the advantages of using birds as pathology-free model of type-2 diabetes (T2DM). Using this new perspective, we observed that birds are missing the RAGE gene, considered an important factor in the development of diabetic complications. In this paper, we identify two additional Maillard-reaction-related characteristics of birds that may, in part, account for their ability to cope successfully with chronic hyperglycemia. A. Compared to mammals, blood plasma of birds has significantly higher concentrations of taurine and other free amino acids that act as scavengers of reactive carbonyls. B. There are also indications that avian blood-plasma contains lower concentrations of methylglyoxal (MG) due, in part, to its decreased production by avian erythrocytes. Our deductions are based on relatively meager experimental data and are therefore somewhat speculative. One certain outcome of our study, however, is the idea that birds are a productive venue for the study of Maillard reactions and etiology of diabetic complications. We anticipate that results of such studies will support the hypothesis identifying methylglyoxal as a key intermediate in the development of diabetic complications. If methylglyoxal is indeed such a central player, then prevention and control of diabetic complications may become transformed into more a circumscribed and tractable problem whose goals will be to minimize the production of MG and to maximize its elimination by detoxification or by scavenging.Rejuvenation Research 03/2014; · 2.92 Impact Factor