Inducible and Constitutive Formation of -Fructofuranosidase (Inulase) in Batch and Continuous Cultures of the Yeast Kluyveromyces fragilis

Abstract

Inulase production by Kluyveromyces fragilis on various fermentable and non-fermentable carbon sources was examined in carbon-limited continuous culture. Fructose and sucrose supported superior inulase yields [above 24 pmol sucrose hydrolysed min-l (mg cell dry wt)-l at pH 5.0, 50 "C], while some other carbon sources, including lactose, galactose, ethanol and lactate, did not stimulate inulase formation beyond basal levels. Thus fructose was identified as the primary physiological inducer. Isolation of a constitutive mutant also provided genetic evidence for the inducible nature of inulase in the wild-type. The mutant was generated spontaneously and selected in continuous culture. It produced high inulase activities in continuous culture irrespective of the carbon source. Inulase formation in the wild-type and mutant strain was further controlled by general carbon catabolite repression as suggested by enzyme yield patterns in batch and continuous culture.

Database mining and transcriptional analysis of genes encoding inulin-modifying enzymes of Aspergillus niger

“…In the yeast Kluyveromyces fragilis, fructose has been described as an inducer of inulinase expression (Grootwassink & Hewitt, 1983). The results shown in Fig.…”

“…At this point we can not rule out the possibility that also these lower concentrations of sucrose might have induced expression of the inulinolytic enzymes at earlier time points. Assuming that the mRNAs might not be very stable, we might have missed the induction as we have analysed the expression only after 4 h. In an identical experimental set-up, also fructose was tested as an inducer, previously reported to act as an inducer for inulinase expression in the yeast K. lactis (Grootwassink & Hewitt, 1983). We obtained no evidence that fructose acted as an inducer for the expression of the inulinolytic enzymes in A. niger.…”

“…Fructose has been shown to induce expression of inulinase in the yeast K. fragilis (Grootwassink & Hewitt, 1983). The expression analysis of the inulinolytic genes in the wild-type A. niger strain and the DcreA strain after growth on fructose did not result in detectable expression of any of the genes, indicating that fructose is not the inducing molecule for expression (Fig.…”

Induction and metabolite regulation of levanase synthesis in Bacillus subtilis

“…The corresponding mutations were mapped on the chromosome of B. subtilis by PBS1 transduction between leuA and aroD (16) and were located near the structural gene for levanase (21). The sacL mutations specifically affect levanase synthesis and have no effect on the other saccharolytic enzymes, sucrase and levansucrase (16).Previous reports indicate that levanase synthesis in sacL mutants is repressed by the presence of rapidly catabolized carbon sources (16), a property common to most inulases and invertases (8,13,14,17). The expression of inulase by Kluyveromyces fragilis (13) and S. mutans (14) is inducible by fructose and seems to be regulated by a dual control * Corresponding author.…”

“…Such inulase or levanase activities have been found in yeasts (13,34), in filamentous fungi (34), and also in some bacteria, among them Streptococcus mutans (3,5,35), Clostridium acetobutylicum (8), and B. subtilis (16).…”