Page 84 - The Biochemistry of Inorganic Polyphosphates

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Enzymes of polyphosphate biosynthesis and degradation
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from GTP. For ppk1, the activity of PolyP synthesis is 4-fold higher than the activity of
PolyP utilization. Secondly, ppk2 uses GTP and ATP equally well in PolyP synthesis, but
ppk1 is strictly specific for ATP in the PolyP synthesis. PolyPs of 15–700 phosphate residues
can serve as a substrate, but PolyPs of 30–50 residues are optimal for GTP synthesis by
ppk2. GDP is more preferable than ADP among nucleoside diphosphate acceptors. Thus,
in P. aeruginosa the ppk2 function is first of all utilization of PolyP.
The gene encoding ppk2 (ppk2) was identified from the amino acid sequence of the
purified protein. It encodes a protein of 357 amino acids with a molecular mass of 40.8
kDa. Both of the polyphosphate kinases ppk1 and ppk2 may be involved in regulation
of the level of ribonucleoside triphosphates and deoxyribonucleoside triphosphates that
modulate cell division and survival in the stationary phase (Ishige et al., 2002).
Sequences homologous to ppk2 were found in two other proteins in P. aeruginosa,intwo
Archaea, and in 32 other bacteria including several pathogenic species (Ishige et al., 2002;
Zhang et al., 2002). Table 6.1 shows the distribution of ppk1 and ppk2 gene homologues
in microorganisms (Zhang et al., 2002). These are lacking in most currently sequenced
genomes of Eukarya and Archaea (Kornberg et al., 1999).
Polyphosphate kinase activity was observed in the yeast vacuolar membrane, but the
activity when consuming PolyP and forming ATP was higher than in PolyP synthesis
(Shabalin et al., 1977). The enzyme has not been purified, and the significance of PolyP
synthesisusingATPincertainmembranefractionsinyeastcellsneedsfurtherinvestigations.
The level of this activity observed in yeast was insufficient to explain the synthesis of a
large amount of PolyP in this microorganism.
The polyphosphate kinase from the yeast cell homogenate purified by Felter and Stahl
4
1

(Felter and Stahl, 1973) was shown to be actually diadenosine-5 ,5 -P ,P tetraphosphate

a,b-phosphorylase (AP 4 phosphorylase). The enzyme acting in concert with one or more
yeast polyphosphatases provided the production of 32 P-labelled ATP in the presence of
32 P-labelled PolyP and ADP (Booth and Guidotti, 1995):
32
32
[ P] PolyP −−−−−−−−−→ [ P] PolyP n − 1 + 32 P i (6.2)
n
exopolyphosphatase
1
4

diadenosine-5 , 5 -P , P tetraphosphate −−−−−−−−→ 2 ADP (6.3)
AP 4 phosphorylase
32 32
[ P i ] + ADP −−→ [ P] ADP + P i (6.4)
32
32
[ P] ADP + ADP −−−−−−−→ [ P] ATP + AMP (6.5)
adenylate kinase
The resulting reaction was probably observed by Felter and Stahl (1973):
32
32
32
[ P] PolyP + ADP −−→ [ P] PolyP + [ P] ATP (6.6)
n n − 1
It was reported that the archaeon Sulfolobus acidocaldarius possessed a glycogen-bound
polyphosphate kinase, which was active only as a native complex with glycogen (Skorko
et al., 1989). This result is doubted by Cardona et al. (2001) who repeated the purification