Identification, cloning, and heterologous expression of a mammalian fructosamine-3-kinase

• Published in: Diabetes (New York, N.Y.) Vol. 49; no. 10; pp. 1627 - 1634
• Main Authors: DELPIERRE, Ghislain, RIDER, Mark H, COLLARD, Francois, STROOBANT, Vincent, VANSTAPEL, Florent, SANTOS, Helena, VAN SCHAFTINGEN, Emile
• Format: Journal Article
• Language: English
• Published: Alexandria, VA American Diabetes Association 01-10-2000
• Subjects: Adenosine Triphosphate - pharmacology; Amino Acid Sequence; Animals; Base Sequence; Biological and medical sciences; Chromatography; Cloning, Molecular; Complications and side effects; Diabetes; Diabetes mellitus; Diabetes. Impaired glucose tolerance; DNA, Complementary - chemistry; Electrophoresis, Polyacrylamide Gel; Endocrine pancreas. Apud cells (diseases); Endocrinopathies; Enzymes; Erythrocytes - enzymology; Etiopathogenesis. Screening. Investigations. Target tissue resistance; Fructose - analogs & derivatives; Fructose - metabolism; Gene Expression; Humans; Kinases; Magnetic Resonance Spectroscopy; Medical sciences; Mice; Molecular Sequence Data; Morpholines - metabolism; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor) - blood; Phosphotransferases (Alcohol Group Acceptor) - genetics; Physiological aspects; Proteins; Sequence Alignment; Transfection; Endocrinopathy; Characterization; Human; Blood cell; Enzyme; Protein kinase; Transferases; Diabetes mellitus; Red blood cell; Complication; Molecular cloning
• DOI: 10.2337/diabetes.49.10.1627
• EISSN: 1939-327X
• ISSN: 0012-1797

Identification, cloning, and heterologous expression of a mammalian fructosamine-3-kinase.
G Delpierre ,
M H Rider ,
F Collard ,
V Stroobant ,
F Vanstapel ,
H Santos and
E Van Schaftingen
Laboratory of Physiological Chemistry, Université Catholique de Louvain, Brussels, Belgium.
Abstract
Fructosamines are thought to play an important role in the development of diabetic complications. Little is known about reactions
that could metabolize these compounds in mammalian tissues, except for recent indications that they can be converted to fructosamine
3-phosphates. The purpose of the present work was to identify and characterize the enzyme responsible for this conversion.
Erythrocyte extracts were found to catalyze the ATP-dependent phosphorylation of 1-deoxy-1-morpholinofructose (DMF), a synthetic
fructosamine. The enzyme responsible for this conversion was purified approximately 2,500-fold by chromatography on Blue Sepharose,
Q Sepharose, and Sephacryl S-200 and shown to copurify with a 35,000-M(r) protein. Partial sequences of tryptic peptides were
derived from the protein by nanoelectrospray-ionization mass spectrometry, which allowed for the identification of the corresponding
human and mouse cDNAs. Both cDNAs encode proteins of 309 amino acids, showing 89% identity with each other and homologous
to proteins of unknown function predicted from the sequences of several bacterial genomes. Both proteins were expressed in
Escherichia coli and purified. They were shown to catalyze the phosphorylation of DMF, fructoselysine, fructoseglycine, and
fructose in order of decreasing affinity. They also phosphorylated glycated lysozyme, though not unmodified lysozyme. Nuclear
magnetic resonance analysis of phosphorylated DMF and phosphorylated fructoseglycine showed that the phosphate was bound to
the third carbon of the 1-deoxyfructose moiety. The physiological function of fructosamine-3-kinase may be to initiate a process
leading to the deglycation of fructoselysine and of glycated proteins.