A simple colorimetric assay for measuring fructosamine 3 kinase activity

Area of Science:

• Biochemistry
• Enzymology
• Clinical Chemistry

Background:

• Fructosamine 3 kinase (FN3K) is a deglycating enzyme with potential to mitigate diabetes-induced organ damage by removing glucose from glycated proteins.
• Developing a straightforward colorimetric method for assaying FN3K activity in human body fluids is crucial for further research.

Purpose of the Study:

• To develop and validate a simple colorimetric assay for measuring fructosamine 3 kinase (FN3K) activity in human serum.
• To establish reference values for FN3K activity in a healthy adult population.
• To investigate the relationship between FN3K activity, diabetes, and specific genetic polymorphisms.

Main Methods:

• Glycated bovine serum albumin (BSA) was prepared and used as a substrate for FN3K.
• Serum samples were incubated with the substrate, MgCl2, and ATP.
• Fructosamine concentration was measured before and after incubation to determine FN3K activity.
• FN3K and ferroportin single nucleotide polymorphisms (SNPs) were genotyped.

Main Results:

• A reliable colorimetric assay for FN3K activity in human serum was established with a within-assay coefficient of variation (CV) of 6.0%.
• Reference values for serum FN3K activity were determined (14.2±1.6 U/L) and found to be independent of age and sex.
• Serum FN3K activity in diabetic patients was comparable to controls, but erythrocyte FN3K activity was significantly higher. A weak but significant correlation was observed between FN3K activity and HbA1c.
• FN3K activity showed dependence on ferroportin Q248H genotypes.

Conclusions:

• A simple colorimetric method for determining FN3K activity in human serum has been successfully developed.
• This assay holds promise for investigating the role of deglycation processes in the context of diabetes mellitus.