Processing of PDGF gene products determines interactions with glycosaminoglycans.
The platelet derived growth factor (PDGF), a mitogen for mesenchymal cells, may be bound to and inhibited by heparin and other glycosaminoglycans. PDGF is a homo- or heterodimer of A- and B-chains. They occur as short (A109 and B110) and long (A125 and B160) isoforms. The latter contain basic carboxyl-terminal extensions. Dimeric A125 binds to heparin through its basic extension in a two-step reaction. The mechanism involves a conformational change and is consistent with a Monod-Wyman-Changeux allosteric model. Previous indirect experiments suggested that three critical amino acids (basic R111, K116 and polar T125) might be involved. Here, direct binding experiments using dimeric full-length mutants in surface plasmon resonanse analysis showed that all three critical amino acids in an R(X)4K(X)8T-motif contributed in a concerted manner to the high affinity binding. Mutations of these amino acids to alanine resulted in large thermodynamic changes, loss of the allosteric mechanism and order(s) of magnitude lower binding affinity. The binding mechanism and affinity of long dimeric rB were similar to the mutants. Short dimeric rA109 and rB110 showed 100 times lower binding affinity than rA125. Consequently, interactions with glycosaminoglycans in tissues varies between PDGF isoforms and may influence their local accumulation and activity.
The Wallenberg Laboratory for Cardiovascular Research, Sahlgrenska University Hospital, S-413 45 Göteborg, Sweden. Florentina.Lustig@wlab.wall.gu.se
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Lustig
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Hoebeke
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Simonson
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Ostergren-Lundén
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Bondjers
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Rüetchi
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Fager
D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D019281:Dimerization; D006025:Glycosaminoglycans / Q000378:metabolism*; D006493:Heparin / Q000378:metabolism; D006495:Heparin, Low-Molecular-Weight / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D010982:Platelet-Derived Growth Factor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020349:Surface Plasmon Resonance; D013696:Temperature; D013816:Thermodynamics
D006025:Glycosaminoglycans; D006495:Heparin, Low-Molecular-Weight; D010982:Platelet-Derived Growth Factor; D011993:Recombinant Fusion Proteins; D006493:Heparin