Study On Enzyme Mimics For Degradation Of Fructosyl-Lysine Based On Polypetide-Cu²⁺ Complex

Non-enzymatic glycosylation in human body is a complex chemical reaction process of reducing sugar and amino acids and proteins.The increase and accumulation of AGEs,the final products of non-enzymatic glycosylation,in human body are associated with many diseases,such as diabetes mellitus and atherosclerosis.Fructosamine is a relatively stable product of the non-enzymatic glycosylation process and can be oxidized and degraded by fructosamine oxidase,which is why fructosamine oxidase is a potential tool for protein desugation and treatment of related diseases,as well as fructosamine oxidase has used to detect fructosamine in human serum to reflect average blood glucose levels over 2-3 weeks.However,the preparation method of fructosamine oxidase is complicated and the production cost is high,which seriously restricts its further research and application,so it is important to develop an enzyme mimics with catalytic activity of fructosamine oxidase for fructosamine degradation.Herein,an enzyme mimics based on peptide and copper ions was constructed to degrade small molecule fructosamine.In this work,a small molecule fructosamine-fructosyl-lysine was synthesized as a substrate for studies of enzyme mimics and characterized by NMR hydrogen spectroscopy and high-resolution mass spectrometry.Next,a series of polypeptides rich in histidine were synthesized by solid-phase synthesis and characterized by matrix assisted laser ionizing flight time mass spectrometry.Synthetic polypeptides and copper ions were self-assembled in neutral buffer,resulting in sheet morphology of different sizes and both had β-sheet secondary structures.The longer the peptide chain length,the larger the size of the sheet structure but the self-assembly of individual histidine and copper ions had no secondary structures.Fructosyl-lysine was degraded by a complex of polypeptide and copper ion selfassembly,and it was concluded that the catalytic activity of the self-assembled complex of polypeptide and copper ions was significantly higher than that of copper ions alone and it was speculated that after copper ions and polypeptide were coordinated,the redox potential of copper ions was adjusted to improve its catalytic activity.The catalytic activity of mimic enzymes increased with the growth of peptide chain under the same conditions.However,the complex of histidine and copper ions has little catalytic activity,which may be related to the failure of self-assembly of histidine and copper ions to form suitable coordination domains.The catalytic activity of the enzyme mimics was higher than 25 ℃ at 37 ℃ under the same conditions.At 37 ℃,the catalytic activity of the enzyme mimics was highest,at 17.83%,at a 10:1 concentration of polypeptide H13 and copper ions.In addition,we found that polypeptide H13 and zinc,nickel and iron ions can self-assemble into sheets,granules and spheres with sizes of 100 μm,100 nm and 20 μm in neutral buffers,respectively,and all haveβ-sheet secondary structures.At 37 ℃,the self-assembled complex formed by zinc ion,nickel ion,iron ion and polypeptide H13 showed little catalytic activity toward oxidation of the substrate fructosyl-lysine,the results showed that the redox potential of copper ions was an important factor in the catalytic effect.