Abstract
Autotrophic conversion of CO2 to value-added biochemicals has received considerable attention as a sustainable route to replace fossil fuels. Particularly, anaerobic acetogenic bacteria are naturally capable of reducing CO2 or CO to various metabolites. To fully utilize their biosynthetic potential, an understanding of acetogenesis-related genes and their regulatory elements is required. Here, we completed the genome sequence of the syngas fermenting Eubacterium limosum ATCC 8486 and determined its transcription start sites (TSS). We constructed a 4.4 Mb long circular genome with a GC content of 47.2% and 4,090 protein encoding genes. To understand the transcriptional and translational regulation, the primary transcriptome was augmented, identifying 1,458 TSSs containing a high pyrimidine (T/C) and purine nucleotide (A/G) content at the −1 and +1 position, respectively, along with 1,253 5′-untranslated regions, and principal promoter elements such as −10 (TATAAT) and −35 (TTGACA), and Shine-Dalgarno motifs (GGAGR). Further analysis revealed 93 non-coding RNAs, including one for potential transcriptional regulation of the hydrogenase complex via interaction with molybdenum or tungsten cofactors, which in turn controls formate dehydrogenase activity of the initial step of Wood-Ljungdahl pathway. Our results provide comprehensive genomic information for strain engineering to enhance the syngas fermenting capacity of acetogenic bacteria.
Highlights
Feature Genome size G + C content Genes Predicted protein encoding sequences Predicted sequences encoding RNA genes Coding density ribosomal RNAs (rRNA) transfer RNAs (tRNA) Genes with function prediction Genes assigned to Clusters of Orthologous Groups (COG) Genes with Pfam domains Genes with signal peptides
Acetogenic bacteria inhabit diverse environments and culture conditions vary, indicating that the regulatory elements embedded in their genomes may differ across strains
Genes encoding the methyl and carbonyl branches of the WLP were located in two different genomic regions, whereas a single gene cluster encodes both methyl and carbonyl branch in other acetogenic Clostridia[21]
Summary
Feature Genome size (base pairs) G + C content Genes Predicted protein encoding sequences Predicted sequences encoding RNA genes Coding density rRNA tRNA Genes with function prediction Genes assigned to COGs Genes with Pfam domains Genes with signal peptides. To understand the regulation of syngas fermentation by acetogenic bacteria, it is important to comparatively analyse the genomic features among acetogenic bacteria and determine the genome embedded non-coding regulatory elements, such as promoters, 5′UTRs, and non-coding RNAs (ncRNA), which play key roles in transcriptional and post-transcriptional regulation. We analysed the complete genomic sequence and determined the genome-wide TSSs of E. limosum ATCC 8486. To this end, we integrated a long-read sequencing and short-read sequencing platform to obtain an accurate genomic sequence, and identified the metabolic pathways responsible for syngas fermentation. Based on the integrated analysis with the complete genomic sequence, we suggest that the regulatory features such as principal promoter elements and 5′UTRs orchestrate the WLP and energy conservation system of E. limosum
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