Resolving the effects of albumin glycation using the quartz crystal microbalance

G.T Heller,M.H Sazinsky,T.J Zwang,M.S Johal

doi:10.1051/matecconf/20130402001

Abstract

Both lysine and arginine residues are particularly important at receptor sites for binding anionic ligands. These receptor sites may become compromised via non-enzymatic glycation. While lysine residues are glycated in the presence of glucose, arginine residues are predominantly glycated by α-oxoaldehydes like glyoxal. This study used a quartz crystal microbalance with dissipation monitoring (QCM-D) to examine the binding affinity of surface immobilized human serum albumin (HSA) to hemin after the HSA was pre-incubated with glucose or glyoxal. We found it necessary to pre-expose the HSA functionalized crystal surface to hemin to block irreversible unintended interactions. Glycation with glucose showed little affect on HSA’s affinity for hemin, however, modification with glyoxal showed diminished hemin binding capacity. Despite the hemin-blocking step, we were unable to obtain Kd values consistent with those in literature, which we attribute to other unaccounted for nonspecific interactions. This study highlights the need for a kinetic QCM-D analysis method that accounts for unintended interactions at the sensor surface so that the hemin-blocking step may be eliminated.

Highlights

Several investigations have reported a link between poor glycemic control in diabetics and increased serum iron levels, suggesting a correlation between non-enzymatic glycation and increased serum iron [1,2,3,4]
While lysine residues are glycated on human serum albumin (HSA) in the presence of glucose [6], arginine residues are predominantly glycated in the presence of α-oxoaldehydes like glyoxal [7], a biproduct of sugar autoxidation that is more prevalent in diabetics than the general population
HSA incubated with glyoxal had a diminished hemin binding capacity, suggesting that modification of HSA by dicarbonyls in diabetics may result in increased concentrations of free hemin available to bacteria

Summary

Introduction

Several investigations have reported a link between poor glycemic control in diabetics and increased serum iron levels, suggesting a correlation between non-enzymatic glycation and increased serum iron [1,2,3,4]. While lysine residues are glycated on human serum albumin (HSA) in the presence of glucose [6], arginine residues are predominantly glycated in the presence of α-oxoaldehydes like glyoxal [7], a biproduct of sugar autoxidation that is more prevalent in diabetics than the general population. Arginine and lysine residues are both commonly found at anionic ligand binding pockets of proteins. Due to the prevalence of arginine residues at this location, it is likely that glycation with glyoxal or other sugars could result in a decrease in affinity of HSA for hemin, an iron containing porphyrin. We hypothesize that the glycation of HSA by sugars known to glycate at arginine residues disrupts HSA’s ability to function in iron homeostasis, which would promote bacterial infection by increasing the availability of serum iron

Methods

Results

Conclusion