Purification of feline lysosomal α-mannosidase, determination of its cDNA sequence and identification of a mutation causing α-mannosidosis in Persian cats

Abstract

alpha-Mannosidosis is a lysosomal storage disorder that is caused by the deficiency of lysosomal alpha-mannosidase. Feline alpha-mannosidosis is a well-characterized animal model used for studying pathological and therapeutic aspects of lysosomal storage disorders. We here report the purification of feline liver lysosomal alpha-mannosidase and determination of its cDNA sequence. The active enzyme consisted of three polypeptides, with molecular masses of 72, 41 and 12 kDa, joined by non-covalent forces. The cDNA sequence of feline lysosomal alpha-mannosidase was determined from reverse transcriptase PCR products obtained from skin fibroblast mRNA. The deduced amino acid sequence contained the N-terminal sequences of the 72 and 41 kDa peptides. This indicated that the enzyme is synthesized as a single-chain precursor with a putative signal peptide of 50 amino acids followed by a polypeptide chain of 957 amino acids, which is cleaved into the three polypeptides of the mature enzyme. The deduced amino acid sequence was 81.1 and 83.2% identical with the human and bovine lysosomal alpha-mannosidases sequences respectively. A 4 bp deletion was identified in an alpha-mannosidosis-affected Persian cat by DNA sequencing of reverse transcriptase PCR products. The deletion resulted in a frame shift from codon 583 and premature termination at codon 645. No lysosomal alpha-mannosidase activity could be detected in the liver of this cat. A domestic long-haired cat expressing a milder alpha-mannosidosis phenotype than the Persian cat had a lysosomal alpha-mannosidase activity of 2% of normal. This domestic long-haired cat did not possess the 4 bp deletion, proving molecular heterogeneity for feline alpha-mannosidosis.

Characterization of a Human Core-specific Lysosomal α1,6-Mannosidase Involved in N-Glycan Catabolism

“…An enzyme deficiency in LysMan leads to the lysosomal storage disease, ␣-mannosidosis, characterized by glycan accumulation and pathology in humans (13,14), cattle (15), cats (16,17), mice (18) and guinea pigs (19). ␣-Mannosidosis models in various mammalian species reveal a remarkable variation in the glycan structures that accumulate.…”

A Radiation Hybrid Map of the Cat Genome: Implications for Comparative Mapping

“…The domestic cat has long played a role in comparative medicine beginning with comparative anatomy, physiology, and biochemistry, extending to the present, in which the species provides numerous models for human heritable and infectious disease (http://www.angis.org.au/Databases/BIRX/omia/) including lysosomal storage diseases (Yogalingam et al 1996;Berg et al 1997;Fyfe et al 1999), polycystic kidney disease (Biller 1996), and AIDS (FIV-feline immunodeficiency virus; Willett et al 1997). The domestic cat also provides a valuable reference for comparative genome analysis, displaying remarkable syntenic conservation relative to the human genome (O'Brien et al 1997a(O'Brien et al ,b, 1999Rettenberger et al 1995;Wienberg et al 1997).…”

Apparent Diffusion Coefficient Reveals Gray and White Matter Disease, and T2 Mapping Detects White Matter Disease in the Brain in Feline Alpha-Mannosidosis

“…Peripheral blood leukocytes were tested at 1 day of age for the 4-base-pair deletion causing AMD, by using described methods. 23 All affected cats were homozygous for the mutation, and all normal cats were null for the mutation. Cats were sedated with intravenous ketamine (2.2 mg/kg) and acepromazine maleate (0.1 mg/kg) and were given intravenous atropine sulfate (0.02 mg/kg).…”