Involvement of the carboxy-terminal propeptide of β-glucuronidase in its compartmentalization within the endoplasmic reticulum as determined by a synthetic peptide approach

The proenzyme form of β-glucuronidase is compartmentalized in large quantities within the endoplasmic reticulum by binding to the esterase, egasyn. Also, the propeptide of the proenzyme form of β-glucuronidase is likely located at the carboxyl terminus. We have, therefore, tested if this carboxyl-terminal peptide is important in binding to egasyn. A polyclonal antibody to a 30-mer synthetic peptide, corresponding to the carboxyl-terminal 30 amino acids of pro-β-glucuronidase, provided evidence that egasyn binds to the carboxyl terminus of β-glucuronidase. This antibody interacted with proenzyme β-glucuronidase-egasyn complexes in which one, two, or three egasyn molecules were bound to the β-glucuronidase tetramer, but not with those complexes (M₄) which contained four egasyn molecules. We interpret these results as indicating that all available carboxyl termini of the β-glucuronidase proenzyme tetramer are shielded by egasyn in the M₄ complexes. The same antibody did not recognize the mature lysosomal form of β-glucuronidase, indicating that only the proenzyme form of microsomal β-glucuronidase contains the original carboxyl terminus. Also, the synthetic 30-mer was found to be a specific and potent inhibitor (50% inhibition at 1.3 μM) of the esterase activity of purified egasyn but exhibited little inhibitory activity toward other purified esterases including a rat trifluoroacetylated esterase or egasyn esterase from another species. Together, these data describe a potent interaction of the exposed carboxyl terminus of precursor glucuronidase with the esterase catalytic site of egasyn, which in turn results in the specific localization of glucuronidase within the lumen of the endoplasmic reticulum.