Page 91 - The Biochemistry of Inorganic Polyphosphates

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Enzymes of polyphosphate degradation 75
have been a gradual transition from PolyP to ATP as the phosphoryl donor in glucose
phosphorylation (Phillips et al., 1999).
6.2.2 NAD Kinase (ATP:NAD 2 -Phosphotransferase,

EC 2.7.1.23)
This enzyme catalyses the following reaction:

ATP + NAD −−→ ADP + NADP (6.10)
The above reaction was known many years ago (Kornberg, 1950; Wang and Ka-
plan, 1954) and found both in procaryotes and eucaryotes. In Brevibacterium ammoni-
agenes (Murata et al., 1979), Micrococcus luteus and Corynebacterium ammoniagenes
(Fillipovich et al., 2000) phosphorylation of NAD using PolyP as a phosphate donor was
revealed:
PolyP + NAD −−→ PolyP + NADP (6.11)
n n−1
It has been established that in some bacteria one enzyme displays both activities (Kawai
et al., 2000). An enzyme with both PolyP- and ATP-dependent NAD kinase activities
was isolated from Micrococcus flavus. This enzyme is a dimer consisting of 34 kDa sub-
units. A gene Rv1695 has been found in Mycobacterium tuberculosis and proposed to also
be a PolyP-dependent NAD kinase. By cloning and expression in E. coli, Rv1695 was
shown to encode PolyP/ATP–NAD kinase and was named as ppnk. The ppnk product, a
recombinant PolyP/ATP–NAD kinase (Ppnk), was purified and characterized. This enzyme
was a tetramer consisting of 35 kDa sub-units when expressed in E. coli. PolyP/ATP–NAD
kinases of M. flavus and Ppnk of M. tuberculosis H37Rv phosphorylated NAD, using PolyP
and nucleoside triphosphates as the phosphoryl donors (Kawai et al., 2000).
NAD kinase was purified to homogeneity from E. coli. The enzyme was a hexamer con-
sisting of 30 kDa sub-units and utilized ATP or other nucleoside triphosphates as phosphoryl
donors for the phosphorylation of NAD. This enzyme could not use PolyP. The deduced
amino acid sequence exhibited a homology with that of M. tuberculosis PolyP/ATP–NAD
kinase (Kawai et al., 2001). Therefore, NAD kinases show the same features as PolyP/ATP
glucose kinases. The enzymes are active or not active with PolyP, depending on the mi-
croorganism under study. The evolutionally older Mycobacteria possess both activities, in
contrast to the evolutionally younger E. coli. This fact, together with data on the distribution
of polyphosphate glucokinase in bacteria, confirms the idea of the greater role of PolyPs in
cell energetics at the earliest stages of evolution (Kulaev, 1971, 1974).

6.2.3 Exopolyphosphatase (Polyphosphate
Phosphohydrolase, EC 3.6.1.11)
One of the most important enzymes involved in PolyP metabolism is exopolyphosphatase,
the enzyme that splits P i from the end of the PolyP chain:

PolyP + H 2 O −−→ PolyP n − 1 + P i (6.12)
n