Solution Structure of Heparin-binding Growth Factor Midkine - Interaction with Heparin

 Upon the signal transduction of growth factors, growth factors bind to the extracellular domain of receptors, causing the oligomerization of receptors and intermolecular autophosphorylation of tyrosine residues that result in stimulation of protein tyrosine kinase activity. Therefore, receptor oligomerization is the key step for the signal transduction of growth factors. In the case of heparin-binding growth factors, heparin sulfate proteoglycans are required for the oligomerization of ligands and/or the presentation of these oligomers to their appropriate signaling receptors.

Midkine (MK) is a heparin-binding growth factor discovered by Muramatsu et al, (refer to review 1). MK enhances neurite outgrowth, neuronal cell survival and plasminogen activator activity. Strong expression of MK is found in various human tumors. Therefore, MK is expected to be a tumor marker and target for tumor therapy. The amino acid sequence of MK is unrelated to other proteins and forms a new growth factor family with pleiotrophin, which was discovered after MK.

MK is structurally divided into two domains. We determined the solution structures of the two domains by NMR (2): MK residues 1-59 (N-domain) and MK residues 62-104 (C-domain). Both domains have anti-parallel -sheet structures (Fig. 1). Most of the heparin-binding activity is located on the C-domain. In the C-domain, basic residues on the -sheet and those on the long loop make clusters on the same surface. These clusters are identified as the heparin-binding sites by NMR and site-directed mutagenesis. Although N-domain also has many basic residues, its heparin-binding activity is low compared to that of the C-domain. This is because the basic residues of the N-domain do not form clusters suitable for interacting with the sulfate groups on heparin oligosaccharides like C-domain.
Fig. 1 Solution structures of the N-domain and C-domain (2).
Main chain folds of each peptide from the N- to C-terminus are represented by gradation from white to green or pink. The disulfide bonds are shown in yellow.
It has been reported that some MK activity requires dimerization of MK. The binding of MK to heparin of more than 20 monosaccharide units induced MK dimerization (2). These results suggest that the MK dimer is the active form, and that heparan sulfate is required for the dimerization of MK. We proposed a molecular model for the dimer formation of MK on heparin oligosaccharides. If C-domains form a head-to-head dimer, the shape and the location of the basic clusters and those of the sulfate clusters on heparin oligosaccharide fit well with each other (Fig. 2). This feature implies that the MK dimer binds heparin more strongly than the monomer.
Fig. 2 The model for the C-domain dimer (blue, basic residues; red, acidic residues; green, Gln) and heparin (pink, oxygens of sulfate groups).
Positively charged clusters (blue circles) of the C-domain fit to negatively charged clusters (pink circles) of heparin.
MK receptors are thought to be proteoglycans such as protein-tyrosine phosphatase (PTP) and syndecan, and members of the low density lipoprotein receptor-related protein (LRP) family. Oligomerization of MK induced by heparan sulfate would be the first key step in signal transduction. Considering the examples of other growth factors, astructural biological approach is expected to be helpful in elucidating visually the signal transduction mechanism of MK.
Wakana Iwasaki (RIKEN Harima Institute)
Fuyuhiko Inagaki (Graduate School of Pharmaceutical Sciences, Hokkaido University)
References (1) Muramatsu T: Midkine and pleiotrophin: two related proteins involved in development, survival, inflammation and tumorigenesis. J. Biochem. 132, 359-371, 2002
(2) Iwasaki W, Nagata K, Hatanaka H, Inui T, Kimura T, Muramatsu T, Yoshida K., Tasumi M, Inagaki F: Solution structure of midkine, a new heparin-binding growth factor. EMBO J. 16, 6936-6946, 1997
PG-A01 Growth Factor and Heparan Sulfate(Masayuki Ishihara)
GG-A02 Functions of syndecan-4 (Takashi Muramatsu and Tetsuhito Kojima)
Sep. 30, 2004

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