Question

1) how many polypeptide chains does PFK-1 consist of?

2) is it a homodimer, heterodimer, homotrimer, heterotrimer, homotetramer or heterotetramer? and how did you come to this conclusion?

3) What is involved in the folding and unfolding of a polypeptide chain? are hydrogen bonds formed and broken or covalent bonds and are they just formed/broken in the protein backbone or from one protein strand to another?

4) what kind of interaction is the interaction between one of the ADP sugar ring hydroxyl groups and a peptide backbone carbonyl? (pi-stacking, dipole-dipole, ion-ion, hydrophobic interaction and/or hydrogen bonding)

5) what interaction is found between the F6P phosphate group binding to the protein via interactions with 3 arginine sidechains? (pi-stacking, dipole-dipole, ion-ion, hydrophobic interaction and/or hydrogen bonding)

Answer 1

1) Phosphofructokinase-1 (PFK-1) is a glycolytic enzyme that catalyzes the transfer of a phosphoryl group from ATP to fructose-6-phosphate (F6P) to yield ADP and fructose-1,6-bisphosphate (FBP). It contains 8 polypeptide chains.

2) Mammalian PFK1 is a 340 kd tetramer composed of different combinations of three types of subunits: muscle (M), liver (L), and platelet (P). The composition of the PFK1 tetramer differs according to the tissue type it is present in. For example, mature muscle expresses only the M isozyme, therefore, the muscle PFK1 is composed solely of homotetramers of M4. The liver and kidneys express predominantly the L isoform. In erythrocytes, both M and L subunits randomly tetramerize to form M4, L4 and the three hybrid forms of the enzyme (ML3, M2L2, M3L). As a result, the kinetic and regulatory properties of the various isoenzymes pools are dependent on subunit composition. Tissue-specific changes in PFK activity and isoenzymic content contribute significantly to the diversities of glycolytic and gluconeogenic rates which have been observed for different tissues.^[3]

PFK1 is an allosteric enzyme and has a structure similar to that of hemoglobin in so far as it is a dimer of a dimer. One half of each dimer contains the ATP binding site whereas the other half the substrate (fructose-6-phosphate or (F6P)) binding site as well as a separate allosteric binding site. Each subunit of the tetramer is 319 amino acids and consists of two domains: one that binds the substrate ATP, and the other that binds fructose-6-phosphate. Each domain is a b barrel, and has cylindrical b sheet surrounded by alpha helices.

On the opposite side of the each subunit from each active site is the allosteric site, at the interface between subunits in the dimer. ATP and AMP compete for this site. The N-terminal domain has a catalytic role binding the ATP, and the C-terminal has a regulatory role