Characterization of polypeptides serologically and structurally related to hexosaminidase in cultured fibroblasts

Abstract

Human fibroblasts synthesize several polypeptides that assort into the various forms of hexosaminidase (hex). We report here the occurrence of three newly identified, hexosaminidase-related polypeptides resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of immunoprecipitates from [35S]methionine-labeled cell extracts. These polypeptides, called band 2 (75,000), band 3 (70,000), and band 4 (63,000), were immunoprecipitated by an antiserum specific to placental hex I2. They are distinct from pre-α- (60,000) and pre-β- (58,000) precursor polypeptides and the α- (56,000), β(a)- (27,000), and β(b)- (27,000) polypeptides of the mature hex A (αβ(a)β(b)) and hex B (2[β(a)β(b)]). When fibroblast extracts were chromatographed on DEAE-Sepharose, bands 2, 3, and 4 were eluted together in fractions before hex A, in a position characteristic of serum and placental hex I2 and serum hex P. This suggests that bands 2, 3, and 4 might represent the polypeptides of a fibroblast hex I. The analysis of partial proteolytic digests of the radioactively labeled polypeptides revealed that bands 2 amd 3, pre-β, β(a) had several peptides in common, suggesting that they are structurally related to each other. However, bands 2, 3, and 4 were present in extracts of Tay-Sachs (pre-α and α deficiency) and Sandhoff cells (pre-β, β(a), and β(b) deficiency) and appeared later than pre-β in pulse-chase experiments. These results suggest that bands 2 and 3 occur independently of pre-β and β(a) and are probably specified by different mRNA, whether from the same gene or distinct but homologous genes.