Cloning, expression and chromosomal localization of a human testis 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene

6-Phosphofructo-2-kinase/fructose 2,6-bisphosphatase (PFK-2/FBPase-2) is a bifunctional enzyme responsible for the synthesis and breakdown of Fru-2,6- P-2, a key metabolite in the regulation of glycolysis. Several genes encode distinct PFK-2/FBPase-2 isozymes that differ in their tissue distribution and enzyme regulation. In this paper, we present the isolation of a cDNA fr om a human testis cDNA library that encodes a PFK-2/FBPase-2 isozyme. Seque ncing data show an open reading frame of 1407 nucleotides that codifies for a protein of 469 amino acids. This has a calculated molecular weight of 54 kDa and 97% similarity with rat testis PFK-2/FBPase-2, with complete conse rvation of the amino acid residues involved in the catalytic mechanism. Flu orescence in-situ hybridization (FISH) localized testis PFK-2/FBPase-2 gene (PFKFB4) in human chromosome 3 at bands p21-p22. A Northern blot analysis of different rat tissues showed the presence of a 2.4-kb mRNA expressed spe cifically in testis. In mammalian COS-1 cells, the human testis cDNA drives expression of an isozyme with a molecular weight of 55 kDa. This isozyme s hows clear PFK-2 activity. Taken together, these results provide evidence f or a new PFK-2/FBPase-2 gene coding for a human testis isozyme. (C) 1999 El sevier Science B.V. All rights reserved.