Plant Nucleotide Metabolism. Hiroshi Ashihara

apt5 AT5g11160 HGPRT HX G X PRPP Jerusalem artichoke^a) 7.5–8.5 6.1 5.5 8.8 Le Floc'h and Lafleuriel (1981) A. thaliana ^b) 176 29 ^c) hpt/gpt AT1G71750 Liu et al. (2007)

      A, Adenine; PRPP, 5-phosphoribosyl-1-pyrophosphate. Cytokinin bases: BA, benzyladenine; iP, isopentenyladenine; tZ, trans-zeatin. HX, Hypoxanthine; G, guanine; X, xanthine.

      —, not determined.

a) Native enzyme.

b) Recombinant enzyme.

c) Activity was detected but no data was published on the Km value.

Compared with APRT, only a few kinetic analyses of HGPRT activity have been carried out with plant enzymes (Table 5.2). A native HGPRT, partially purified from Jerusalem artichoke shoots, exhibited the low Km values for hypoxanthine, guanine, and PRPP (<10 μM), which have high affinities for the enzyme. The rate of IMP formation from hypoxanthine was greater than GMP production from guanine. Enzyme activity was inhibited by the products, IMP and GMP (Le Floc'h and Lafleuriel 1981).

      Molecular and functional analysis of HGPRT of A. thaliana was carried out by Liu et al. (2007). The kinetic analysis of the recombinant HGPRT revealed that the enzyme catalyses the conversion of guanine and hypoxanthine to their respective nucleoside monophosphates, but that xanthine is not a substrate. The Km values of the recombinant enzyme for hypoxanthine and guanine were 176 and 29 μM, respectively. The relatively low affinity of the A. thaliana recombinant enzyme for hypoxanthine is different to that from other sources, including humans (Keough et al. 1999) and yeast (Ali and Sloan 1982), as well as the native plant enzyme (Le Floc'h and Lafleuriel 1981), which displays similar Km values (<10 μM) towards guanine and hypoxanthine. The reason why the recombinant A. thaliana HGPRT possesses a substantially higher affinity for guanine than for hypoxanthine remains to be resolved. Like the enzymes in humans and yeast, A. thaliana HGPRT does not act on xanthine (Liu et al. 2007).

      5.3.3 Xanthine Phosphoribosyltransferase

      As noted above, in most prokaryotes and nearly all eukaryotes, HGPRT catalyses the salvage of hypoxanthine and guanine. However, in a few species, xanthine salvage enzyme activity is also detected. Guanine and xanthine phosphoribosyltransferase (GXPRT, EC 2.4.2.22) activity has been observed in Lactobacillus casei and E. coli (Krenitsky et al. 1970). More specific xanthine phosphoribosyltransferase (XPRT) is found in Leishmania donovani, a protozoan parasite. This enzyme preferentially uses xanthine; the Km value for xanthine (7 μM) is much lower than for hypoxanthine and guanine (100–450 μM) (Jardim et al. 1999). In plants, low phosphoribosylation of xanthine was detected in tea leaf extracts in vitro (Deng and Ashihara 2010). However, it is unclear whether this activity is due to a distinct XPRT or if it is a side reaction of HGPRT (Ashihara et al. 2018).

5.4 Properties of Nucleoside Kinases

      Nucleoside kinases are enzymes which catalyse the transfer of γ-phosphate from ATP to nucleosides leading to formation of nucleoside-5′-monophosphates.

      5.4.1 Adenosine Kinase

Adenosine kinase (AK, ATP: adenosine-5′-phosphotransferase, EC 2.7.1.20) catalyses the phosphorylation of adenosine to AMP using ATP as the phosphate donor (reaction 2 in Figure 5.2). The enzyme was first isolated from yeast and animal tissues (Caputto, 1951; Kornberg and Pricer, 1951). AK is widely distributed in plant cells and tissues and there are reports on both purified native and recombinant enzymes (Table 5.3).

Table 5.3 Properties of the native and recombinant adenosine kinase (AK) and inosine/guanosine kinase (IGK) from plants.

				Km values (μM)
Enzyme	Enzyme source	Isozyme	Optimum pH	AR	ATP	tZR	iPR	DHZR	Gene	TAIR Locus	References
AK	Wheat germ		6.8–7.2	8.7			31				Chen and Eckert (1977)
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