Journal of Comparative Physiology B

, Volume 160, Issue 3, pp 309–316 | Cite as

cGMP-stimulated protein kinase phosphorylates pyruvate kinase in an anoxia-tolerant marine mollusc

  • Stephen P. J. Brooks
  • Kenneth B. Storey
Article

Summary

A cystosolic protein kinase that phosphorylates pyruvate kinase (PK) in vitro has been identified in crude homogenates of heart, radular retractor, and foot muscle from the anoxia-tolerant marine whelk Busycon canaliculatum. Protein kinase action was measured by following changes in PK kinetic parameters: phosphorylated PK has a higher Kmvalue for phosphoenolpyruvate and a lower I50 value for l-alanine. The crude protein kinase readily phosphorylated PK in a Mg2+-and ATP-dependent manner in the absence of any added effector. This activity was not affected by the addition of either cAMP (a stimulator of protein kinase A) or Ca2+ plus phorbol 12-myristate 13-acetate (stimulators of protein kinase C) to the incubation medium. Addition of cGMP to the homogenate, however, increased the rate of PK phosphorylation giving a 3–4-fold increase in the rate of change in PK kinetic parameters that was readily apparent after 5h. Complete time-courses of changes in PK kinetic parameters in the presence and absence of cGMP showed that cGMP increased the rate, but not the final extent, of PK phosphorylation. These results indicate that PK inactivation by enzyme phosphorylation in response to anoxia in whelk tissues may be mediated by a cyclic GMP stimulated protein kinase in response to changing levels of cGMP. This conclusion was further supported by data indicating that the total activity of protein kinase was the same in both anoxic and aerobic animals, and that the total PK phosphatase activity was also constant. Changes in PK phosphorylation during anoxia are not, therefore, the result of changes in the total amount of protein kinase or phosphatase.

Key words

Anaerobiosis Pyruvate kinase cGMP-dependent protein kinase Marine molluscs Reversible phosphorylation control of enzymes 

Abbreviations

cAMP

adenosine 3′:5′-monophosphate

cGMP

guanosine 3′:5′-monophosphate

PK

pyruvate kinase

PMA

phorbol 12-myristate, 13-acetate

PEP

phosphoenolpyruvate

Km

Michaelis constant

I50

inhibitor concentration that reduces enzyme activity by 50%

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Copyright information

© Springer-Verlag 1990

Authors and Affiliations

  • Stephen P. J. Brooks
    • 1
    • 2
  • Kenneth B. Storey
    • 1
    • 2
  1. 1.Institute of BiochemistryCarleton UniversityOttawaCanada
  2. 2.Department of BiologyCarleton UniversityOttawaCanada

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