Molecular cloning and characterization of murine and human N-acetylglucosamine kinase

N-Acetylglucosamine is produced by the endogenous degradation of glycoconju gates and by the degradation of dietary glycoconjugates by glycosidases. It enters the pathways of aminosugar metabolism by the action of N-acetylgluc osamine kinase. In this study we report the isolation and characterization of a cDNA clone encoding the murine enzyme. An open reading frame of 1029 b ase pairs encodes 343 amino acids with a predicted molecular mass of 37.3 k Da. The deduced amino-acid sequence contains matches of the sequences of ei ght peptides derived from tryptic cleavage of rat N-acetylglucosamine kinas e. The recombinant murine enzyme was functionally expressed in Escherichia coli BL21 cells, where it displays N-acetylglucosamine kinase activity as w ell as N-acetylmannosamine kinase activity. The complete cDNA sequence of human N-acetylglucosamine kinase was derived from the nucleotide sequences of several expressed sequence tags. An open r eading frame of 1032 base pairs encodes 344 amino acids and a protein with a predicted molecular mass of 37.4 kDa. Similarities between human and muri ne N-acetylglucosamine kinase were 86.6% on the nucleotide level and 91.6% on the amino-acid level. Amino-acid sequences of murine and human N-acetylg lucosamine kinase show sequence similarities to other sugar kinases, and al l five sequence motifs necessary for the binding of ATP by sugar kinases ar e present. Tissue distribution of murine N-acetylglucosamine kinase revealed an ubiqui tous occurrence of the enzyme and a very high expression in testis. The siz e of the murine mRNA was 1.35 kb in all tissues investigated, with the exce ption of testis, where it was 1.45 kb mRNA of the murine enzyme was continu ously expressed during mouse development. mRNA of the human enzyme was expr essed in all investigated human tissues, as well as in cancer cell lines. I n both the tissues and the cancer cell lines, the human mRNA was 1.35 kb in size.