Mapping of glycolytic enzyme-binding sites on human erythrocyte band 3.

Previous work has shown that GAPDH (glyceraldehyde-3-phosphate dehydrogenase), aldolase, PFK (phosphofructokinase), PK (pyruvate kinase) and LDH (lactate dehydrogenase) assemble into a GE (glycolytic enzyme) complex on the inner surface of the human erythrocyte membrane. In an effort to define the molecular architecture of this complex, we have undertaken to localize the binding sites of these enzymes more accurately. We report that: (i) a major aldolase-binding site on the erythrocyte membrane is located within N-terminal residues 1-23 of band 3 and that both consensus sequences D6DYED10 and E19EYED23 are necessary to form a single enzyme-binding site; (ii) GAPDH has two tandem binding sites on band 3, located in residues 1-11 and residues 12-23 respectively; (iii) a PFK-binding site resides between residues 12 and 23 of band 3; (iv) no GEs bind to the third consensus sequence (residues D902EYDE906) at the C-terminus of band 3; and (v) the LDH- and PK-binding sites on the erythrocyte membrane do not reside on band 3. Taken together, these results argue that band 3 provides a nucleation site for the GE complex on the human erythrocyte membrane and that other components near band 3 must also participate in organizing the enzyme complex.

[1]  F. Clarke,et al.  Identification of an actin binding region in aldolase , 1993, FEBS letters.

[2]  Samuel E. Lux,et al.  Blood: Principles and Practice of Hematology , 1995 .

[3]  J. Hoffman.  ATP compartmentation in human erythrocytes , 1997, Current opinion in hematology.

[4]  P. Low,et al.  Characterization of the reversible conformational equilibrium of the cytoplasmic domain of erythrocyte membrane band 3. , 1984, The Journal of biological chemistry.

[5]  H. Knull,et al.  A glycolytic enzyme binding domain on tubulin. , 1997, Archives of biochemistry and biophysics.

[6]  S. Rapoport,et al.  A reappraisal of the binding of cytosolic enzymes to erythrocyte membranes. , 1989, Trends in biochemical sciences.

[7]  R. Moriyama,et al.  Partial Characterization of the Cytoplasmic Domain of Human Kidney Band 3 (*) , 1995, The Journal of Biological Chemistry.

[8]  H. Kliman,et al.  Glyceraldehyde-3-phosphate dehydrogenase of rat erythrocytes has no membrane component. , 1985, Biochimica et biophysica acta.

[9]  P. Low,et al.  Tyrosine phosphorylation of band 3 inhibits peripheral protein binding. , 1987, The Journal of biological chemistry.

[10]  M. Tanner,et al.  A band 3-based macrocomplex of integral and peripheral proteins in the RBC membrane. , 2003, Blood.

[11]  Philip S Low,et al.  Assembly and regulation of a glycolytic enzyme complex on the human erythrocyte membrane. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[12]  P. Low,et al.  Identification of a Critical Ankyrin-binding Loop on the Cytoplasmic Domain of Erythrocyte Membrane Band 3 by Crystal Structure Analysis and Site-directed Mutagenesis* , 2003, The Journal of Biological Chemistry.

[13]  C. Buscaglia,et al.  Characterization of an Aldolase-binding Site in the Wiskott-Aldrich Syndrome Protein* , 2006, Journal of Biological Chemistry.

[14]  R. Reithmeier,et al.  Identification of the carbonic anhydrase II binding site in the Cl(-)/HCO(3)(-) anion exchanger AE1. , 2000, Biochemistry.

[15]  H. Lardy,et al.  PURIFICATION AND SOME PROPERTIES OF RABBIT SKELETAL MUSCLE PHOSPHOFRUCTOKINASE. , 1965, The Journal of biological chemistry.

[16]  L. Ercolani,et al.  Colocalization of GAPDH and band 3 (AE1) proteins in rat erythrocytes and kidney intercalated cell membranes. , 1992, The American journal of physiology.

[17]  P. Low,et al.  The N-terminal 11 amino acids of human erythrocyte band 3 are critical for aldolase binding and protein phosphorylation: implications for band 3 function. , 2005, Blood.

[18]  I. Tsai,et al.  Effect of red cell membrane binding on the catalytic activity of glyceraldehyde-3-phosphate dehydrogenase. , 1982, The Journal of biological chemistry.

[19]  P. Low,et al.  Regulation of glycolysis via reversible enzyme binding to the membrane protein, band 3. , 1993, The Journal of biological chemistry.

[20]  P. Low,et al.  Partial structural characterization of the cytoplasmic domain of the erythrocyte membrane protein, band 3. , 1981, The Journal of biological chemistry.

[21]  G. T. Rich,et al.  Glyceraldehyde-3-phosphate dehydrogenase release from erythrocytes during haemolysis. No evidence for substantial binding of the enzyme to the membrane in the intact cell. , 1984, The Biochemical journal.

[22]  R. Kaul,et al.  The aldolase-binding site of the human erythrocyte membrane is at the NH2 terminus of band 3. , 1981, The Journal of biological chemistry.

[23]  A. Saltiel,et al.  Aldolase Mediates the Association of F-actin with the Insulin-responsive Glucose Transporter GLUT4* , 1999, The Journal of Biological Chemistry.

[24]  T. Steck,et al.  Mode of interaction of phosphofructokinase with the erythrocyte membrane. , 1985, The Journal of biological chemistry.

[25]  K. Uyeda,et al.  The interaction of phosphofructokinase with erythrocyte membranes. , 1979, The Journal of biological chemistry.

[26]  H. Lodish,et al.  Cloning and characterization of band 3, the human erythrocyte anion-exchange protein (AE1). , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[27]  Association of phosphofructokinase and aldolase with the membrane of the intact erythrocyte. , 1984, The Journal of biological chemistry.

[28]  D. Wallach,et al.  Electrophoretic analysis of the major polypeptides of the human erythrocyte membrane. , 1971, Biochemistry.

[29]  P. Low,et al.  Crystallographic structure and functional interpretation of the cytoplasmic domain of erythrocyte membrane band 3. , 2000, Blood.

[30]  P. Vincent,et al.  Blood — Principles and Practice of Hematology , 1995 .

[31]  G. T. Rich,et al.  Lack of binding of glyceraldehyde-3-phosphate dehydrogenase to erythrocyte membranes under in vivo conditions. , 1985, Biochimica et biophysica acta.

[32]  T. Steck,et al.  Interaction of the aldolase and the membrane of human erythrocytes. , 1977, Biochemistry.

[33]  E. Tisdale,et al.  Glyceraldehyde-3-phosphate Dehydrogenase Interacts with Rab2 and Plays an Essential Role in Endoplasmic Reticulum to Golgi Transport Exclusive of Its Glycolytic Activity* , 2004, Journal of Biological Chemistry.

[34]  D. Yannoukakos,et al.  Phosphorylation sites in human erythrocyte band 3 protein. , 1991, Biochimica et biophysica acta.

[35]  D. Schubert,et al.  The complex of band 3 protein of the human erythrocyte membrane and glyceraldehyde-3-phosphate dehydrogenase: stoichiometry and competition by aldolase. , 2002, Biochimica et biophysica acta.

[36]  M. Fautsch,et al.  The identification of myocilin-associated proteins in the human trabecular meshwork. , 2006, Experimental eye research.

[37]  P. Low,et al.  Expression, purification, and characterization of the functional dimeric cytoplasmic domain of human erythrocyte band 3 in Escherichia coli , 1992, Protein science : a publication of the Protein Society.

[38]  R. Morrow.  A study into human erythrocyte membrane protein association. , 2003 .

[39]  Hans Ulrich Bergmeyer,et al.  Methods of Enzymatic Analysis , 2019 .

[40]  G. Somero,et al.  Properties of the interaction between phosphofructokinase and actin. , 1989, Archives of biochemistry and biophysics.