Caprine beta-mannosidase: sequencing and characterization of the cDNA and identification of the molecular defect of caprine beta-mannosidosis.

The complete sequence of the caprine beta-mannosidase cDNA coding region has been determined, and a mutation that is associated with caprine beta-mannosidosis has been identified. Reverse transcriptase-polymerase chain reactions were performed using primers based on bovine and, later, goat cDNA sequences to produce an overlapping series of amplicons covering the entire coding region. The composite cDNA codes for an 879-amino-acid peptide that has four potential N-glycosylation sites. Comparison of the caprine and bovine cDNAs reveals that 96.3% of the nucleotides and 95.2% of the deduced amino acids are identical. A single-base deletion at position 1398 of the coding sequence was identified in the cDNA isolated from a goat affected with beta-mannosidosis. This deletion results in a shift in the reading frame and a premature termination of translation, yielding a deduced peptide of 481 amino acids. An assay, developed to determine the presence or absence of this mutation, confirmed that animals affected with beta-mannosidosis were homozygous for the mutation and that obligate carriers in a caprine beta-mannosidosis colony were heterozygous. This assay accurately distinguished between mutation carrier and noncarrier goats and was used for prenatal diagnosis using DNA collected from fetal fluids. The assay also confirmed chimerism in a goat with an atypically mild beta-mannosidosis phenotype. Thus, this application enables assessment of the efficacy of engraftment of hematopoietic stem cells after prenatal transfer from donor sources.