The substrate dependence and product inhibition of three different fructokinases and three different hexokinases from growing potato (
Solanum tuberosum L.) tubers was investigated. The tubers contained three specific fructokinases (FK1, FK2, FK3) which had a high affinity for fructose
K
m=64, 90 and 100 (M) and effectively no activity with glucose or other hexose sugars. The affinity for ATP (
K
m=26, 25 and 240 M) was at least tenfold higher than for other nucleoside triphosphates. All three fructokinases showed product inhibition by high fructose (
K
i=5.7, 6.0 and 21 mM) and were also inhibited by ADP competitively to ATP. Sensitivity to ADP was increased in the presence of high fructose, or fructose-6-phosphate. In certain conditions, the
K
i (ADP) was about threefold below the
K
m (ATP). All three fructokinase were also inhibited by fructose-6-phosphate acting non-competitively to fructose (
K
i=1.3 mM for FK2). FK1 and FK2 showed very similar kinetic properties whereas FK3, which is only present at low activities in the tuber but high activities in the leaf, had a generally lower affinity for ATP, and lower sensitivity to inhibition by ADP and fructose. The tuber also contained three hexokinases (HK1, HK2, HK3) which had a high affinity for glucose (
K
m=41, 130 and 35 M) and mannose but a poor affinity for fructose (
K
m=11, 22 and 9 mM). All three hexokinases had a tenfold higher affinity for ATP (
K
m=90, 280 and 560 M) than for other nucleoside triphosphates. HK1 and HK2 were both inhibited by ADP (
K
i=40 and 108 M) acting competitively to ATP. HK1, but not HK2, was inhibited by glucose-6-phosphate, which acted non-competitively to glucose (
K
i=4.1 mM). HK1 and HK2 differed, in that HK1 had a narrower pH optimum, a higher affinity for its substrate, and showed inhibition by glucose-6-phosphate. The relevance of these properties for the regulation of hexose metabolism in vivo is discussed.Abbreviations FK
fructokinase
- Fru6P
fructose-6-phosphate
- Glc6P
glucose-6-phosphate
- HK
hexokinase
- NTP
nucleoside triphosphate
- P
i
inorganic phosphate
- UDPGlc
uridine-5-diphosphoglucose
This work was supported by the Deutsche Froschungsgemeinschaft (SFB 137). We are grateful to Professor E. Beck (Lehrstuhl für Pflanzenphysiologie, Universität Bayreuth, FRG) for providing laboratory facilities.
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