J. Biol. Chem., Vol. 263, Issue 32, 16631-16636, 11, 1988
Further enzymatic characteristics of a thylakoid protein kinase
S Coughlan, J Kieleczawa and G Hind
Biology Department, Brookhaven National Laboratory, Upton, New York 11973.
The enzymatic characteristics of a protein kinase purified from thylakoids
are further described. ATP (KM approximately 30 microM) and Mg2+ ion
(greater than 1.0 mM) were required for activity, while ADP was a
competitive inhibitor (Ki = 100 microM). Activity was 55% inhibited by the
sulfhydryl inhibitor p-chloromercuribenzoate (1 mM) and was less sensitive
to substituted maleimides. Lysine-rich histones (H1) were utilized as an
exogenous phosphorylation substrate both by thylakoid-bound kinase and by
isolated enzyme; threonine was predominantly phosphorylated by the in situ
enzyme, whereas the isolated enzyme phosphorylated closely related serine
residues as determined by peptide mapping. Detergents that proved useful in
extracting the kinase from thylakoids markedly inhibited activity of the
isolated enzyme, whereas Triton X-100 and 3-[(3-
cholamidopropyl)dimethylammonio]-1-propane-sulfonic acid had little effect.
The enzyme could be freed from detergent and behaved as an active monomer
on size-exclusion chromatography. The phosphate contents of the
light-harvesting chlorophyll a/b protein complex of photosystem II isolated
from maximally phosphorylated thylakoid membranes of spinach and pea were
equivalent to approximately 6% and approximately 19% phosphorylation,
respectively. Corresponding values for nonphosphorylated membranes were
approximately 3% and approximately 14.5%.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
