(A) Alignment of vertebrate midkine proteins. Identical residues are in black, conservative substitutions in gray. Conserved cysteins are indicated by squares, residues important for heparin binding by triangles (cluster I) or circles (cluster II). An arginine residue essential for receptor binding is marked by an asterisk; glutamine residues Q43 and Q95 important for dimerization are marked by arrows. Accession nos. as follows: Danio rerio Mdka AF503614; Oncorhynchus mykiss Mdka CA346000 (EST); Salmo salar BG934205 (EST); Ictalurus punctatus BM495405 (EST); Danio rerio Mdkb NM_131716; Oncorhynchus mykiss Mdkb CA365321 (EST); Takifugu rubripes scaffold S004868 (http://fugu.hgmp.mrc.ac.uk); Tetraodon nigroviridis see Methods; Oryzias latipes BJ523610 (EST); Homo sapiens AAA58478; Mus musculus AAH12244; Gallus gallus P24052; Xenopus laevis BAA07658.
(B) Both zebrafish midkine genes encode secreted proteins. Western blot detection of MYC-epitope-tagged Mdka and Mdkb proteins in supernatants of human 293 kidney fibroblasts transfected with pCS-mdka-MYC and pCS-mdkb-MYC. A HA-tagged version of Mdkb was cotransfected as a control.
