Molecular cloning, structure, promoters and regulatory elements for transcription of the Bacillus licheniformis encoded regulon for xylose utilization

Abstract

In this article we describe the cloning of the xyl regulon encoding xylose utilization from Bacillus licheniformis by complementation of a xyl mutant of B. subtilis. The xylose isomerase encoding gene, xylA, was sequenced and identified by its extensive homology to other xylose isomerases. The expression of xylA is regulated on the level of transcription by a repressor protein encoded by xylR. Its gene has the opposite orientation of xylA and the start codons are 181 bp apart. A deletion of xylR renders xylA expression constitutive. The xylR sequence was determined and is discussed with respect to its homology to other xylR structures. Primer extension analyses of the xylA and xylR transcripts under repressing and including conditions define their promoters and confirm the regulation of xylA transcription. Furthermore, some induction of the xylR transcript by xylose is also observed. The regulatory sequence of both genes consists of a bipolar promoter system and contains three palindromic sequence elements. Their potential functions with respect to xylA and xylR regulation are discussed. The primary structures of the genes, promoters and regulatory sequences are compared to the xyl regulons encoded by B. subtilis, B. megaterium, Staphylococcus xylosus and E. coli. Homology is greatest between the B. subtilis and B. megaterium encoded xyl genes while the B. licheniformis borne genes are clearly more distant. The next greater differences are found to the S. xylosus and the greatest to the E. coli encoded genes. These results are discussed with respect to the taxonomic relations of these bacteria.

Isolation and characterization of a xylose-dependent promoter from Caulobacter crescentus

“…A third pathway has been found in some pseudomonads; in this case, xylose is first oxidized by xylose dehydrogenase to xylonate, which is then converted by xylonate dehydrase to 3-deoxypentulosonic acid (8). Genes coding for components of the first pathway (xylA, xylose isomerase; xylB, xylulose kinase; xylE, low-affinity uptake; and xylT, high-affinity xylose uptake system) have been cloned from a number of organisms (3,28,34,41,44,45). Genes encoding xylose reductase (XYL1) and xylitol dehydrogenase (XYL2) have also been characterized (1,20).…”

Genetic Analysis of a Novel Pathway for d -Xylose Metabolism in Caulobacter crescentus

“…In the Dahms pathway (7), this compound is converted by an aldolase to pyruvate and glycoaldehyde. In an alternative reaction first demonstrated by Weimberg (27) and confirmed by Watanabe et al (25,26), for L-arabinose degradation in A. brasilense, a dehydratase produces ␣-ketoglutarate semialdehyde, which is then oxidized to ␣-ketoglutarate. The genes identified (through mutation) in this work as necessary for growth on D-xylose (Table 1) and the enzymes they encode are shown beside the appropriate reaction.…”

“…Two routes for D-xylose degradation in microorganisms have been described. Numerous bacteria, including Escherichia coli (15), Bacillus species (24,25), and Lactobacillus species (16), use xylose isomerase to convert D-xylose to xylulose, which is then phosphorylated to enter the pentose phosphate pathway. Although some fungi have recently been shown to use this "bacterial" pathway (11), fungi more commonly transform Dxylose into xylitol by using xylose reductase and xylitol dehydrogenase (13).…”

Glucose and glucose-6-phosphate interaction with Xyl repressor proteins from Bacillus spp. may contribute to regulation of xylose utilization

“…The materials, enzymes, media, and many general methods used in this study have been described earlier (3). In particular, media for growth of B. subtilis (7), B. licheniformis (27), and B. megaterium (22) strains have been described. When necessary, we used chloramphenicol (5 g/ ml), erythromycin (25 g/ml), kanamycin (5 g/ml), or tetracycline (10 g/ml) for selection.…”

“…involves isomerization to xylulose and then phosphorylation to xylulose-5-phosphate. The enzymes for this pathway are encoded by xyl operons consisting of at least two genes encoding xylose isomerase (xylA) and xylulo kinase (xylB), as described for Bacillus subtilis (7,9,31,32), Bacillus megaterium (24), and Bacillus licheniformis (27). These operons are negatively regulated by binding of the Xyl repressor (XylR) to xyl operators (xylO) in the absence of the inducer xylose (16,24,27).…”