Complete Genome Sequence of the Industrial Bacterium Ketogulonicigenium vulgare SKV.

Ketogulonicigenium vulgare has been widely used in vitamin C two-step fermentation, which converts l-sorbose to 2-keto-l-gluonic acid. Here, the complete genome of K. vulgare SKV, which performs better fermentation production than K. vulgare Hbe602, is deciphered to understand the key differences in metabolism between K. vulgare strains SKV and Hbe602.

GENOME ANNOUNCEMENT

Ketogulonicigenium vulgare was identified as a member of the Proteobacteria (1). It can convert l-sorbose to 2-keto-l-gulonic acid (2-KGA), the precursor of vitamin C (2). Mono-cultured K. vulgare grows poorly and the Bacillus spp. are usually cocultivated with it to achieve a high 2-KGA yield (3). Previously, we published the genome of K. vulgare Hbe602 (4). Our research showed that combinational expression of sorbose/sorbosone dehydrogenases and cofactor pyrroloquinoline quinone in K. vulgare Hbe602 could enhance 2-KGA production properly (5). It was interesting to remark that industrial K. vulgare performed better 2-KGA production than K. vulgare Hbe602 when they were cocultured with Bacillus thuringiensis Bc601 (6).

K. vulgare strain SKV (Shandong Luwei Pharmaceutical Co., Ltd.) was cultured in 250-ml flasks at 30°C and 250 rpm for 35 h. The seed medium contains 3 g/L beef extract, 3 g/L yeast powder, 3 g/L corn steep liquor, 0.2 g/L MgSO4, 1 g/L KH2PO4, 1 g/L urea, and 10 g/L peptone. The genomic DNA was isolated using the SDS method. The genome of K. vulgare SKV was sequenced by single-molecule real-time (SMRT) technology (Beijing Novogene Bioinformatics Technology Co., Ltd.). SMRT Analysis version 2.3.0 was used to filter low-quality reads, and the filtered reads were assembled to generate one contig without gaps.

The genome of K. vulgare SKV was annotated through the NCBI Prokaryotic Genome Annotation Pipeline (7) and using BLAST (8) against the Kyoto Encyclopedia of Genes and Genomes (KEGG) database (9) and the Clusters of Orthologous Groups (COG) of proteins database (10). The tRNAs and rRNAs were predicted by tRNAscan (11) and RNAmmer (12), respectively. The origin of replication (oriC) and putative DnaA boxes were identified using Ori-Finder (13). GC-Profile was used to identify the GC content variation in DNA sequences (14).

The genome of K. vulgare SKV consists of one circular chromosome (2,764,573 bp) and one circular plasmid (267,949 bp). The sequence difference between K. vulgare SKV and Hbe602 was analyzed using BLAST, and the sequence similarity between the chromosomes of K. vulgare SKV and Hbe602 is more than 99%. Additionally, the plasmid in K. vulgare SKV is almost the same as plasmid 1 in Hbe602. The better ability of 2-KGA production in K. vulgare SKV may be due to the loss of plasmid 2 in Hbe602. Plasmid 2 of K. vulgare Hbe602 encodes 211 proteins, which are mainly related to transport systems, transcriptional regulators, and dehydrogenases. We identified that overexpression of l-sorbosone dehydrogenase (GI: 939479492) in plasmid 2 of K. vulgare Hbe602 produced an obvious byproduct in K. vulgare (15). Compared with K. vulgare Hbe602, the loss of dehydrogenases may lead to the higher 2-KGA production. We hope these findings can provide insight into the metabolism and gene targets for the strain improvement of K. vulgare.

Accession number(s).

The sequence of the K. vulgare SKV genome has been deposited at DDBJ/EMBL/GenBank under the GenBank accession numbers CP016592 (chromosome) and CP016593 (plasmid).