Abstract
The XYL1 gene of the yeast Pichia stipitis has been isolated from a genomic library using a specific cDNA probe, and its nucleotide (nt) sequence has been determined. In the 5′ noncoding region of the P. stipiti XYL1 gene a TATAAA element (known to be necessary for transcription initiation in most yeast genes) is found at nt −81, and two CCAAT recognition motifs (often referred to as the CCAAT box) are present at nt −146 and −106. The XYL1 encodes a polypeptide of 35 927 Da that constitutes a NADH/NADPH-dependent xylose reductase (XR). The enzyme is part of the xylose-xylulose pathway that is absent or only weakly expressed in Saccharomyces cerevisiae. Extensive homology is found to the N terminus of the XR of Pachysolen tannophilus and Candida shehatae. None of the known cofactor binding domains found in many NAD-dependent dehydrogenases are present in the protein. Transformants of S. cerevisiae containing XYL1 of P. stipitis synthesize an active XR. Fusion of XYL1 with the prokaryotic tac promoter leads to a gene that can be expressed in S. cerevisiae and Escherichia coli.
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