Heparan N-sulfatase: cysteine 70 plays a role in the enzyme catalysis and processing |
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Authors: | Daniele A Di Natale P |
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Affiliation: | Department of Biochemistry and Medical Biotechnologies, Medical School, University of Naples Federico II, Via Pansini 5, 80131, Naples, Italy. daniele@cds.unina.it |
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Abstract: | Sulfatases are members of a highly conserved family of enzymes that catalyze the hydrolysis of sulfate ester bonds from a variety of substrates. The functional correlation reflects a high degree of amino acid sequence similarity along the entire length, in particular in the active site where the C(X)PSR consensus sequence is present. Cysteine undergoes an important co- or post-translation modification essential for the accomplishment of catalytic activity: conversion in formylglycine. In this work, the cysteine of heparan N-sulfatase (NS) was replaced either by a serine (C70S) or by a methionine (C70M) using site-directed mutagenesis. C70S and C70M mutant cDNAs were expressed and analyzed in COS cells; both mutations caused a loss of NS activity; however, while C70S showed a normal precursor form undergoing processing to a reduced mature form within the lysosomes, C70M was poorly synthesized and formed a complex with the molecular chaperone immunoglobulin binding protein. |
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