Human red cells protein kinase in normal subjects and patients with
hereditary spherocytosis, sickle cell disease, and autoimmune hemolytic
anemia
E Beutler, E Guinto and C Johnson
A somewhat simplified modification of a previously described method for the
measurement of red cell membrane phosphorylation by ATP has been devised.
Phosphorylation of membranes was linear with time for only 5- 10 min, and
linearity with membrane concentration was observed only when assays were
limited to short incubation times. Protein kinase activity of hereditary
spherocytosis (HS) membranes was found to be normal. However, the average
phosphorylation after 60 min incubation was less in HS membranes than in
normal membranes. Findings similar to those in HS membranes were observed
in sickle cell disease. The Km of red cell protein kinase for ATP is
approximately 10(-5) M. Membrane phosphate binding sites are not saturated
in either HS or normal membranes after 1 hr incubation with ATP.
Approximately 27% of phosphorylating activity is lost after 1 hr incubation
at 37 degrees C. GTP is a very inefficient phosphate donor. Under the
conditions of measurement employed, the enzyme is slightly stimulated by 1
muM cAMP, but is not stimulated by 1 muM cGMP. Dephosphorylation of red
cell membranes after labeling occurs at a similar rate in HS as in normal
membranes. Although a mild abnormally in membrane phosphorylation is
observed in HS, this could not be demonstrated to be due to a decrease in
protein kinase activity or in alterations of its kinetic properties. The
abnormally seen is not specific for HS.
Volume 48,
Issue 6,
pp. 887-898,
12/01/1976
Copyright © 1976 by The American Society of Hematology
