title: Electrochemical sensing system employing fructosamine 6-kinase enables glycated albumin measurement requiring no proteolytic digestion.
abstract: Currently available enzymatic methods for the measurement of glycated proteins utilize fructosyl amino acid/peptide oxidases (FAOXs/FPOXs) as sensing elements. FAOXs/FPOXs oxidize glycated amino acids or glycated dipeptides but they are not able to accept longer glycated peptides or intact glycated proteins as substrates. Therefore, pretreatment via proteolytic digestion is unavoidable with the current enzymatic methods, and there remains a need for simpler measurement methods for glycated proteins. In this study, in order to develop a novel sensing system for glycated albumin (GA), a marker for diabetes, with no requirement for proteolytic digestion, we created an electrochemical sensor based on fructosamine 6-kinase (FN6K) from Escherichia coli. Uniquely, FN6K can react directly with intact GA unlike FAOXs/FPOXs. The concentration of GA in samples was measured using a carbon-printed disposable electrode upon which FN6K as well as two additional enzymes, pyruvate kinase and pyruvate dehydrogenase were overlaid. A clear correlation between the response current and the concentration of GA was observed in the range of 20-100 µM GA, which is suitable for measurement of GA in diluted blood samples from both healthy individuals and patients with diabetes. The sensing system reported here could be applied to point-of-care-testing devices for measurement of glycated proteins.
authors.affiliation: Department of Biotechnology and Life Science, Graduate School of Engineering, Tokyo University of Agriculture and Technology, Tokyo, Japan.
authors.forename: Miho
authors.initials: M
authors.lastname: Kameya
authors.affiliation: Department of Biotechnology and Life Science, Graduate School of Engineering, Tokyo University of Agriculture and Technology, Tokyo, Japan.
authors.forename: Wakako
authors.initials: W
authors.lastname: Tsugawa
authors.affiliation: Department of Biotechnology and Life Science, Graduate School of Engineering, Tokyo University of Agriculture and Technology, Tokyo, Japan.
authors.forename: Mayumi
authors.initials: M
authors.lastname: Yamada-Tajima
authors.affiliation: Department of Biotechnology and Life Science, Graduate School of Engineering, Tokyo University of Agriculture and Technology, Tokyo, Japan.
authors.forename: Mika
authors.initials: M
authors.lastname: Hatada
authors.affiliation: Department of Biotechnology and Life Science, Graduate School of Engineering, Tokyo University of Agriculture and Technology, Tokyo, Japan.
authors.forename: Keita
authors.initials: K
authors.lastname: Suzuki
authors.affiliation: D epartment of Medical Technology, School of Health Sciences, Graduate School of Bionics, Computer and Media Sciences, Tokyo University of Technology, Tokyo, Japan.
authors.forename: Akane
authors.initials: A
authors.lastname: Sakaguchi-Mikami
authors.affiliation: Department of Engineering, Graduate School of Integrated Science and Technology, Shizuoka University, Tokyo, Japan.
authors.forename: Stefano
authors.initials: S
authors.lastname: Ferri
authors.affiliation: Diabetes Research Institute, Mills-Peninsula Health Services, San Mateo, California, USA.
authors.forename: David C
authors.initials: DC
authors.lastname: Klonoff
authors.affiliation: Department of Biotechnology and Life Science, Graduate School of Engineering, Tokyo University of Agriculture and Technology, Tokyo, Japan. sode@cc.tuat.ac.jp.
authors.forename: Koji
authors.initials: K
authors.lastname: Sode
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D003920:Diabetes Mellitus / Q000097:blood; D055664:Electrochemical Techniques / Q000379:methods*; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000737:chemistry; D019270:Fructosamine / Q000737:chemistry; D017127:Glycation End Products, Advanced; D006801:Humans; D010088:Oxidoreductases / Q000737:chemistry; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry*; D059748:Proteolysis; D011770:Pyruvate Kinase / Q000737:chemistry; D012709:Serum Albumin / Q000032:analysis*; D000093423:Glycated Serum Albumin
chemical_list: D029968:Escherichia coli Proteins; D017127:Glycation End Products, Advanced; D012709:Serum Albumin; D019270:Fructosamine; D010088:Oxidoreductases; D017853:Phosphotransferases (Alcohol Group Acceptor); D011770:Pyruvate Kinase; D000093423:Glycated Serum Albumin
keywords: Diabetes; Fructosamine 6-kinase; Glycated albumin; Glycated protein; Proteolytic digestion