Planta

, Volume 167, Issue 1, pp 9–18 | Cite as

The partial purification and characterisation of gibberellin 2β-hydroxylases from seeds of Pisum sativum

  • V. A. Smith
  • J. MacMillan
Article

Abstract

The gibberellin (GA) 2β-hydroxylases in mature and immature seeds of Pisum sativum have been partially purified and characterised. The enzymes are unstable when stored below pH 7.0 or in the absence of a thiol reagent. The optimum assay pH is between 7.4 and 7.8 and activity is dependent upon the presence of α-ketoglutarate, Fe2+ and ascorbate. The 2β-hydroxylase activities for GA1, GA4, GA9 and GA20 are chromatographically inseparable and correspond to a protein of Mr 44000. The rate of GA 2β-hydroxylation varies according to substrate and some evidence indicates that the 2β-hydroxylase activities for GA1 and GA4 and for GA9 and GA20 may reside in different proteins. During pea seed maturation, the specific activity of the enzyme(s) increases dramatically and reaches a maximum at a time when endogenous GA9, GA20, GA29 and GA51 are also at their greatest concentration. This correlation is not the result of substrate induction of enzyme activity. Since the GA 2β-hydroxylases operate at maximal rate at low substrate concentrations they are incapable of rapidly 2β-hydroxylating excessive quantities of (exogenously applied) GA1 or GA20. On the basis of the kinetic parameters of the GA 2β-hydroxylase activities, a generalised model is discussed for the control of the steady-state levels of bioactive hormone under normal physiological conditions.

Key words

Gibberellin (2β-hydroxylases) Oxidative enzyme (plant) Pisum (gibberellin) Seed (gibberellin metabolism) 

Abbreviations

DTE

dithioerythritol

EDTA

ethylenediaminetetraacetic acid

GAn

gibberellin An

HPLC

high-performance liquid chromatography

HSS

high-speed supernatant

LSS

low-speed supernatant

PMSF

phenylmethane sulphonyl fluoride

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References

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Copyright information

© Springer-Verlag 1986

Authors and Affiliations

  • V. A. Smith
    • 1
  • J. MacMillan
    • 1
  1. 1.Agricultural and Food Research Council Research Group, School of ChemistryUniversity of BristolBristolUK

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