Profound Biotinidase Deficiency Caused by a Point Mutation That Creates a Downstream Cryptic 3′ Splice Acceptor Site Within an Exon of the Human Biotinidase Gene

Biotinidase recycles the vitamin biotin from biocytin upon the degradation of the biotin-dependent carboxylases. We have identified a novel point mutation within the biotinidase gene that encodes the signal peptide in two unrelated individuals with profound biotinidase deficiency. Sequence analysis of genomic DNA from these individuals revealed a G to A transition (G100→A) located 57 bases downstream of the authentic splice acceptor site in exon B. Although this mutation predicts a G34S substitution, it also generates a 3′ splice acceptor site. Sequence of the PCR-amplified cDNA from the homozygous child revealed that all the product was shorter than that of normal individuals and was the result of aberrant splicing. The aberrantly spliced transcript lacked 57 bases, including a second in-frame ATG, that encode most of the putative signal peptide and results in an in-frame deletion of 19 amino acids. The mutation results in failure to secrete the aberrant protein into the blood. This is the first reported example in which a point mutation creates a cryptic 3′ splice acceptor site motif that is used preferentially over the upstream authentic splice site. The preferential usage of the downstream splice site is not consistent with the 5′→3′ scanning model, but is consistent with the exon definition model of RNA splicing.