29Penicillium chrysogenum is used as industrial producer of penicillin. We investigated its

Aldose 1-epimerases or mutarotases (EC 5.1.3.3) are catalyzing the interconversion of α- and β-anomers of hemiacetals of aldose sugars such as D-glucose and D-galactose, and are presumed to play an auxiliary role in carbohydrate metabolism as mutarotation occurs spontaneously in watery solutions. The first step in the Leloir pathway of D-galactose breakdown is preceded by accelerated conversion of β-D-galactopyranose into the α-anomer, the substrate of the anomer-specific D-galactose 1-kinase. Here, we identified two putative aldose-1-epimerase genes (galmA and galmB) in the model organism Aspergillus nidulans, and characterized them upon generation of single- and double deletion mutant strains, as well as overexpressing mutants carrying multiple copies of either. Assaying cell-free extracts from the galmB single- and galm double mutants, we observed that the mutarotation hardly exceeded spontaneous anomer conversion, while galmB multicopy strains displayed higher activities than the wild type, increasing with the copy number. When grown on D-galactose in submerged cultures, biomass formation and D-galactose uptake rates in mutants lacking galmB were considerably reduced. None such effects were observed studying galmA deletion mutants, which consistently behave like the wild type. We conclude that GalmB is the physiologically relevant mutarotase for the utilization of D-galactose in A. nidulans.

In this study, we analyzed the expression of the structural genes encoding the five enzymes comprising the Leloir pathway of D-galactose catabolism in the industrial cell factory Penicillium chrysogenum on various carbon sources. The genome of P. chrysogenum contains a putative galactokinase gene at the annotated locus Pc13g10140, the product of which shows strong structural similarity to yeast galactokinase that was expressed on lactose and D-galactose only. The expression profile of the galactose-1-phosphate uridylyl transferase gene at annotated locus Pc15g00140 was essentially similar to that of galactokinase. This is in contrast to the results from other fungi such as Aspergillus nidulans, Trichoderma reesei and A. niger, where the ortholog galactokinase and galactose-1-phosphate uridylyl transferase genes were constitutively expressed. As for the UDP-galactose-4-epimerase encoding gene, five candidates were identified. We could not detect Pc16g12790, Pc21g12170 and Pc20g06140 expression on any of the carbon sources tested, while for the other two loci (Pc21g10370 and Pc18g01080) transcripts were clearly observed under all tested conditions. Like the 4-epimerase specified at locus Pc21g10370, the other two structural Leloir pathway genes -UDP-glucose pyrophosphorylase (Pc21g12790) and phosphoglucomutase (Pc18g01390) -were expressed constitutively at high levels as can be expected from their indispensable function in fungal cell wall formation.