Carbohydrate-binding protein 35 (Mac-2), a laminin-binding lectin, forms functional dimers using cysteine 186.
Carbohydrate-binding protein 35 (CBP35), also known as the macrophage surface antigen Mac-2, is a lactosamine-specific lectin whose extracellular properties include the ability to agglutinate cells and to bind avidly to the basement membrane glycoprotein laminin. Although these and other properties would be facilitated by dimerization of this lectin, previous studies have argued against multimeric forms of this protein. We report here that macrophage CBP35, purified by laminin affinity chromatography, exists as several distinct species (Mr 35,000, 67,000, and 80,000) when analyzed under non-reducing conditions. This unexpected finding prompted us to study the biochemistry of multimerization using recombinant CBP35 (rCBP35). rCBP35 expressed in Escherichia coli forms disulfide-linked homodimers (Mr 67,000). The dimeric form of CBP35 binds to laminin with higher affinity than does monomer and by a lactosamine-dependent mechanism. Site-directed mutagenesis indicated that cysteine 186, the single cysteine residue in CBP35, is required for dimerization. These results raise the possibility that homo- and heterodimeric forms of CBP35 contribute to its postulated functions in cell-matrix interactions and growth regulation.
Laboratory of Cancer Biology, Deaconess Hospital, Boston, Massachusetts 02115.
H J
HJ
Woo
M M
MM
Lotz
J U
JU
Jung
A M
AM
Mercurio
D000595:Amino Acid Sequence; D000818:Animals; D000943:Antigens, Differentiation / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry* / Q000378:metabolism; D037502:Galectin 3; D006388:Hemagglutinins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015151:Immunoblotting; D007797:Laminin / Q000378:metabolism*; D037102:Lectins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed
D000943:Antigens, Differentiation; D002352:Carrier Proteins; D037502:Galectin 3; D006388:Hemagglutinins; D007797:Laminin; D037102:Lectins; D003545:Cysteine