The fungal L-arabinose catabolic pathway

L-arabinose is a major constituent of plant material so that L-arabinose catabolism is of relevance for microorganisms living on decaying plant material but also in biotechnology where cheap raw materials are used. The fungal pathway, which is distinctly different to the bacterial pathway consists of two NADPH-linked reductases and two NAD-linked dehydrogenases and xylulokinase. Genes coding for enzymes of this pathway are known except for L-arabinitol 4-dehydrogenase (EC 1.1.1.12) and L-xylulose reductase (EC 1.1.1.10). We recently identified these two unknown genes of the fungal pathway. The L-arabinitol 4-dehydrogenase was purified from the mould Trichoderma reesei. Amino acid sequence of peptide fragments were obtained from the purified protein and the corresponding gene cloned by using PCR. The L-xylulose reductase was identified by screening a cDNA library of T. reesei. A strain of S. cerevisiae was constructed which contained all genes of the pathway except a gene for the L-xylulose reductase. To this strain we then transformed a T. reesei cDNA library in a yeast expression vector and screened for growth on L-arabinose. We could demonstrate that the whole pathway is active in S. cerevisiae, in a strain where all the genes of this pathway are over-expressed, leading to growth on L-arabinose and ethanol production under anaerobic conditions, however at a very low rate. We discuss the limiting steps of L-arabinose catabolism and show ways to improve it.