The 67-kDa enzymatically inactive alternatively spliced variant of β- galactosidase is identical to the elastin/laminin-binding protein

Abstract

Our previous studies showed immunological and functional similarities, as well as partial sequence homology, between the enzymatically inactive alternatively spliced variant of human β-galactosidase (S-gal) and the 67- kDa elastin/laminin-binding protein (EBP) from sheep. To define the genetic origin of the EBP further, a full-length human S-gal cDNA clone was constructed and subjected to in vitro transcription/translation. The cDNA was also transfected into COS-1 cells and into the EBP-deficient smooth muscle cells (SMC) from sheep ductus arteriosus (DA). In vitro translation yielded an unglycosylated form of the S-gal protein, which immunoreacted with anti- β-galactosidase antibodies and bound to elastin and laminin affinity columns. S-gal cDNA transfections into COS-1 and DA SMC increased expression of a 67-kDa protein that immunolocalized intracellularly and to the cell surface and, when extracted from the cells, bound to elastin. The S-gal- transfected cells displayed increased adherence to elastin-covered dishes, consistent with the cell surface distribution of the newly produced S-gal- encoded protein. Transfection of DA SMC additionally corrected their impaired elastic fiber assembly. These results conclusively identify the 67-kDa splice variant of β-galactosidase as EBP.