PLANT PHYSIOLOGY , Vol 104, Issue 4 1371-1380, Copyright © 1994 by American Society of Plant Biologists
|
ENVIRONMENTAL AND STRESS PHYSIOLOGY |
Differential Influence of ATP on Native Spinach 70-Kilodalton Heat-Shock Cognates
J. V. Anderson, D. W. Haskell and C. L. Guy
Plant Molecular and Cellular Biology Program, Department of Environmental Horticulture, Institute of Food and Agricultural Sciences, University of Florida, Gainesville, FL 32611-0512
A constitutively expressed class of 70-kD heat-shock cognate (HSC70)
proteins from spinach leaf tissue was purified based on their affinity for
ATP-agarose. The affinity-purified spinach proteins were resolved into at
least three different forms on two-dimensional gels. Under native
conditions, and in the absence of ATP, the affinity-purified proteins were
separated into three molecular mass classes by gel-filtration
chromatography; a monomer of 85 kD, a multimer of 280 kD, and a large
molecular mass oligomer of >650 kD. All molecular mass forms contained a
major protein that migrated at 79 kD on sodium dodecyl
sulfate-polyacrylamide gels. N-terminal sequencing of the 79-kD purified
monomer showed the highest homology to the endoplasmic reticulum-luminal
HSC70. Addition of Mg-ATP to monomeric HSC70 did not alter its migration
during gel filtration. Addition of Mg-ATP to the dimer converted it to
monomer and oligomeric forms, whereas the presence of ATP converted a
fraction of the large molecular mass oligomeric form of HSC70 to dimeric
and monomeric forms. Only the large molecular mass oligomeric HSC70 appears
to autophosphorylate in vitro in the presence of [[gamma]-32P]- ATP. Dimers
and monomers can bind ATP by a nonhydrolytic mechanism and undergo a
conformational change in the presence of Mg-ATP. In this paper we discuss
the effects that ATP may have on the regulation of plant HSC70.
