TY - JOUR
T1 - The interaction of phosphofructokinase with erythrocyte membranes
AU - Higashi, T.
AU - Richards, C. S.
AU - Uyeda, K.
N1 - Copyright:
Copyright 2004 Elsevier B.V., All rights reserved.
PY - 1979
Y1 - 1979
N2 - The interaction of muscle phosphofructokinase with human erythrocyte ghost membranes was investigated. Scatchard analysis of the binding reveals only a single class of binding sites numbering approximately 4 x 105 sites/ghost with an association constant of 2 x 107 M-1. Maximum binding is observed below pH 6.8 and is complete within 30 min at 4°, 24°, and 37°C; binding is inhibited by high ionic strength. Studies on the effect of ligands on the binding show that fructose-1,6-P2, Mg2+, and ATP favor binding, while ADP and 2,3-P2-glycerate do not. The nature of the phosphofructokinase binding site on the membrane was investigated using membranes which had been washed with saline, EDTA, and NaOH to remove various peripheral membrane proteins. The results show that the removal of peripheral proteins from the membrane does not diminish the binding capacity of the membranes, suggesting that an integral membrane protein is the binding site. Phosphofructokinase binding is inhibited by aldolase and glyceraldehyde-3-P dehydrogenase, which have been reported to bind to Band 3 protein. Furthermore, both aldolase and glyceraldehyde-3-P dehydrogenase also mediate the dissociation of phosphofructokinase from these membranes; the amount of phosphofructokinase released is proportional to the amount of aldolase bound. Neither spectrin (0.1 mg/ml) nor hemoglobin (0.1 to 50 mg/ml) influences the binding of phosphofructokinase. The results demonstrate that phosphofructokinase binds to a specific site, probably Band 3 protein, on the inner surface of human erythrocyte membranes. A model which explains the relationship of the binding sites of these enzymes is presented.
AB - The interaction of muscle phosphofructokinase with human erythrocyte ghost membranes was investigated. Scatchard analysis of the binding reveals only a single class of binding sites numbering approximately 4 x 105 sites/ghost with an association constant of 2 x 107 M-1. Maximum binding is observed below pH 6.8 and is complete within 30 min at 4°, 24°, and 37°C; binding is inhibited by high ionic strength. Studies on the effect of ligands on the binding show that fructose-1,6-P2, Mg2+, and ATP favor binding, while ADP and 2,3-P2-glycerate do not. The nature of the phosphofructokinase binding site on the membrane was investigated using membranes which had been washed with saline, EDTA, and NaOH to remove various peripheral membrane proteins. The results show that the removal of peripheral proteins from the membrane does not diminish the binding capacity of the membranes, suggesting that an integral membrane protein is the binding site. Phosphofructokinase binding is inhibited by aldolase and glyceraldehyde-3-P dehydrogenase, which have been reported to bind to Band 3 protein. Furthermore, both aldolase and glyceraldehyde-3-P dehydrogenase also mediate the dissociation of phosphofructokinase from these membranes; the amount of phosphofructokinase released is proportional to the amount of aldolase bound. Neither spectrin (0.1 mg/ml) nor hemoglobin (0.1 to 50 mg/ml) influences the binding of phosphofructokinase. The results demonstrate that phosphofructokinase binds to a specific site, probably Band 3 protein, on the inner surface of human erythrocyte membranes. A model which explains the relationship of the binding sites of these enzymes is presented.
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M3 - Article
C2 - 39927
AN - SCOPUS:0018668105
SN - 0021-9258
VL - 254
SP - 9542
EP - 9550
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 19
ER -