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Chimeric phosphofructokinases involving exchange of the N‐ and C‐terminal halves of mammalian isozymes: Implications for ligand binding sites
Author(s) -
Martínez-Costa Oscar H.,
Sánchez-Martínez Cristina,
Sánchez Valentina,
Aragón Juan J.
Publication year - 2007
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2007.05.059
Subject(s) - isozyme , allosteric regulation , phosphofructokinase , fructose , biochemistry , chemistry , enzyme , ligand (biochemistry) , stereochemistry , c terminus , phosphofructokinase 1 , n terminus , phosphofructokinase 2 , peptide sequence , glycolysis , amino acid , receptor , gene
Two phosphofructokinase (PFK) chimeras were constructed by exchanging the N‐ and C‐terminal halves of the mammalian M‐ and C‐type isozymes, to investigate the contribution of each terminus to the catalytic site and the fructose‐2,6‐P 2 /fructose‐1,6‐P 2 allosteric site. The homogeneously‐purified chimeric enzymes organized into tetramers, and exhibited kinetic properties for fructose‐6‐P and MgATP similar to those of the native enzyme that furnished the N‐terminal domain in each case, whereas their fructose‐2,6‐P 2 activatory characteristics coincided with those of the isozyme that provided the C‐terminal half. This reflected the role of each domain in the formation of the corresponding binding site. Grafting the N‐terminus of PFK‐M onto the C‐terminus of the fructose‐1,6‐P 2 insensitive PFK‐C restored transduction of this signal to the catalytic site, which significance is also discussed.