ALPHA-MANNOSIDOSIS - FUNCTIONAL CLONING OF THE LYSOSOMAL ALPHA-MANNOSIDASE CDNA AND IDENTIFICATION OF A MUTATION IN 2 AFFECTED SIBLINGS

alpha-Mannosidosis (MIM 248500) is an autosomal recessive lysosomal st orage disorder resulting from deficient activity of lysosomal alpha-ma nnosidase (LAMAN) (EC 3.2.1.24), The disease is characterized by massi ve intracellular accumulation of mannose-rich oligosaccharides with re sulting mental retardation, hearing loss, immune deficiency and skelet al changes, We report here the purification and characterization of hu man placenta LAMAN, The enzyme is synthesized as a single-chain precur sor which is processed into three glycopeptides of 70, 42 and 15 kDa, The 70 kDa peptide is further partially proteolysed into three more pe ptides that are joined by disulfide bridges, The laman cDNA sequence w as assembled from overlapping fragments obtained by PCR on human fibro blast and human lung cDNA, The deduced amino acid sequence contains a putative signal peptide of 48 amino acids followed by a polypeptide se quence of 962 amino acids, Northern blot analyses revealed a single tr anscript of similar to 3.5 kb present in all tissues examined but at v arying levels. Two affected siblings of Palestinian origin were homozy gous for a mutation that causes a His-->Leu replacement at a position which is conserved among class 2 alpha-mannosidases from several speci es.