Identification and Functional Analysis of the Gene Cluster for l -Arabinose Utilization in Corynebacterium glutamicum

Abstract

Corynebacterium glutamicum ATCC 31831 grew on L-arabinose as the sole carbon source at a specific growth rate that was twice that on D-glucose. The gene cluster responsible for L-arabinose utilization comprised a six-cistron transcriptional unit with a total length of 7.8 kb. Three L-arabinose-catabolizing genes, araA (encoding L-arabinose isomerase), araB (L-ribulokinase), and araD (L-ribulose-5-phosphate 4-epimerase), comprised the araBDA operon, upstream of which three other genes, araR (LacI-type transcriptional regulator), araE (L-arabinose transporter), and galM (putative aldose 1-epimerase), were present in the opposite direction. Inactivation of the araA, araB, or araD gene eliminated growth on L-arabinose, and each of the gene products was functionally homologous to its Escherichia coli counterpart. Moreover, compared to the wild-type strain, an araE disruptant exhibited a >80% decrease in the growth rate at a lower concentration of Larabinose (3.6 g liter Corynebacterium glutamicum is a gram-positive soil microorganism able to secrete large amounts of glutamic acid under suitable conditions (32, 57). Consequently, this organism been used for a long time for industrial production of amino acids and nucleotides, among other primary metabolites (17). Moreover, its potential as a commodity chemical producer is the focus of increasing research efforts (21, 39). Commonly, C. glutamicum is able to grow on various carbon sources, such as hexoses, sugar alcohols, and organic acids (9,29,30). In a future where pentose sugar-rich lignocellulosics should play an important role in global renewable energy, growth of C. glutamicum on pentose sugar-based media is vital to its continued relevance as a producer of commodity chemicals. This has not been possible owing to the lack of essential pentose sugar assimilation pathways in C. glutamicum (4). This contrasts with the characteristics of two major industrial workhorse bacteria, the gram-negative organism Escherichia coli and the grampositive organism Bacillus subtilis, which can grow on L-arabinose as the sole carbon source using the same metabolic pathway. L-Arabinose uptake by E. coli and B. subtilis occurs via two distinct systems involving both a low-affinity H ϩ symporter (encoded by araE) and a high-affinity ATP-dependent transport system (araFGH) (5,18,45). After entering the cell, L-arabinose is sequentially converted to L-ribulose, L-ribulose 5-phosphate, and D-xylulose 5-phosphate by the action of Larabinose isomerase (encoded by araA), L-ribulokinase (araB), and L-ribulose-5-phosphate 4-epimerase (araD), respectively (Fig. 1). D-Xylulose 5-phosphate is eventually metabolized through the reductive pentose phosphate pathway (12). No similar genes for L-arabinose metabolism have been identified in any of the corynebacterial genomes sequenced so far (3,24,38,52,60). Nevertheless, L-arabinose metabolism by C. glutamicum was previously achieved by expression of E. coli araA, araB, and araD genes (26). Although the resulting transformant was able to grow on L-ara...

Characterization of a Novel β-l-Arabinofuranosidase in Bifidobacterium longum

“…Then, HypBA1 (BLLJ_0211) degrades ␤-Ara 2 to L-arabinose. Furthermore, the L-arabinose metabolic enzymes for the conversion to D-xylulose-5-phosphate, which have been characterized in Corynebacterium glutamicum ATCC 31831 (24), exhibit 50 -59% identity with those of B. longum JCM 1217: L-arabinose isomerase (BLLJ_0342), L-ribulokinase (BLLJ_0340), and L-ribulose 5-phosphate 4-epimerase (BLLJ_0341). As a result, HypBA1 plays a key role in B. longum for ␤-L-arabinooligosaccharides usage as a carbohydrate and energy source.…”

AraR, an l -Arabinose-Responsive Transcriptional Regulator in Corynebacterium glutamicum ATCC 31831, Exerts Different Degrees of Repression Depending on the Location of Its Binding Sites within the Three Target Promoter Regions

“…The expression pattern and the location of the galM and araR genes on the chromosome suggest that araR is cotranscribed with galM under the control of the L-arabinose-inducible promoter. However, we also observed upregulation of araR but not of galM in cells grown on L-arabinose compared to cells grown on D-glucose (21). Here, we detected the galM-araR intervening region amplified by RT-PCR performed as described above (Fig.…”

“…The global carbon catabolite repression system, mediated by cAMP in E. coli, has never been found in C. glutamicum. Moreover, it should be noted that C. glutamicum has the ability to consume various carbon sources, including a mixture of D-glucose and L-arabinose, simultaneously (21). Thus, an unidentified unique regulatory system may be involved in the AraR-independent repression of the arabinose catabolic operon in C. glutamicum ATCC 31831.…”

“…Although most C. glutamicum strains are unable to use L-arabinose as a carbon source, C. glutamicum ATCC 31831 has its utilization ability due to the presence of araBDA on its genome (21). We have recently shown that a LacI family transcriptional regulator AraR, which is encoded in the vicinity of the L-arabinose catabolic genes (araBDA), along with its uptake gene (araE) on the genome, binds in vitro to three sites: one in the intergenic region of araB/galM (BS B ) and two upstream of araE (BS E1 /BS E2 ) (Fig.…”

Production of Biofuels from Cellulose of Woody Biomass

“…Corynebacterium glutamicum has also been applied for ethanol production, but it cannot ferment pentoses, unlike E. coli. [58].…”