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-P2, 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 from a human testis cDNA library that encodes a PFK-2/FBPase-2 isozyme. Sequencing 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 54kDa and 97% similarity with rat testis PFK-2/FBPase-2, with complete conservation of the amino acid residues involved in the catalytic mechanism. Fluorescence 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 specifically in testis. In mammalian COS-1 cells, the human testis cDNA drives expression of an isozyme with a molecular weight of 55kDa. This isozyme shows clear PFK-2 activity. Taken together, these results provide evidence for a new PFK-2/FBPase-2 gene coding for a human testis isozyme.
Manzano
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A
Unitat de Bioquímica, Campus de Bellvitge, U.B.C/Feixa Llarga sn E-08907, L'Hospitalet, Spain.
Pérez
J X
JX
Nadal
M
M
Estivill
X
X
Lange
A
A
Bartrons
R
R
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002874:Chromosome Mapping; D002893:Chromosomes, Human, Pair 3; D003001:Cloning, Molecular; D006597:Fructose-Bisphosphatase / Q000235:genetics*; D015870:Gene Expression; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007527:Isoenzymes / Q000235:genetics; D008099:Liver / Q000201:enzymology; D008297:Male; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics*; D009206:Myocardium / Q000201:enzymology; D010732:Phosphofructokinase-1 / Q000235:genetics*; D025481:Phosphofructokinase-2; D012333:RNA, Messenger / Q000378:metabolism; D016415:Sequence Alignment; D013737:Testis / Q000201:enzymology*; D014162:Transfection
D007527:Isoenzymes; D009097:Multienzyme Complexes; D012333:RNA, Messenger; D025481:Phosphofructokinase-2; D010732:Phosphofructokinase-1; D006597:Fructose-Bisphosphatase