Industrial Microorganisms:<i>Corynebacterium glutamicum</i>

Mycobacteria are well known for their taxonomic diversity, their impact on global health, and for their atypical cell wall and envelope. In addition to a cytoplasmic membrane and a peptidoglycan layer, the cell envelope of members of the order Corynebacteriales, which include Mycobacterium tuberculosis, also have an arabinogalactan layer connecting the peptidoglycan to an outer membrane, the so-called “mycomembrane.” This unusual cell envelope composition of mycobacteria is of prime importance for several physiological processes such as protection from external stresses and for virulence. Although there have been recent breakthroughs in the elucidation of the composition and organization of this cell envelope, its evolutionary origin remains a mystery. In this perspectives article, the characteristics of the cell envelope of mycobacteria with respect to other actinobacteria will be dissected through a molecular evolution framework in order to provide a panoramic view of the evolutionary pathways that appear to be at the origin of this unique cell envelope. In combination with a robust molecular phylogeny, we have assembled a gene matrix based on the presence or absence of key determinants of cell envelope biogenesis in the Actinobacteria phylum. We present several evolutionary scenarios regarding the origin of the mycomembrane. In light of the data presented here, we also propose a novel alternative hypothesis whereby the stepwise acquisition of core enzymatic functions may have allowed the sequential remodeling of the external cell membrane during the evolution of Actinobacteria and has led to the unique mycomembrane of slow-growing mycobacteria as we know it today.

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“…), amino acids (e.g., Corynebacterium sp. ), biofuels, and other bioproducts ( Becker and Wittmann, 2016 ; Lewin et al, 2016 ).…”

Section: Introductionmentioning

confidence: 99%

The Mycobacterial Cell Envelope: A Relict From the Past or the Result of Recent Evolution?

Vincent

Nyongesa²,

Morneau³

et al. 2018

Front. Microbiol.

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“…With the development of biotechnology, C. glutamicum has been successfully engineered to serve as a versatile workhorse for industrial bioproduction. Nowadays, this bacterium is used to produce more than 4 million tons of diverse amino acids per year and a wide range of other natural and non-natural products, which are used as feed additives, nutritional supplements, pharmaceutical intermediates, biofuels, and polymer building blocks [ 2 ]. It is estimated that products generated via C. glutamicum fermentation will reach a market size of US$20.4 billion by 2020 [ 3 ].…”

Section: Introductionmentioning

confidence: 99%

Development of a CRISPR/Cas9 genome editing toolbox for Corynebacterium glutamicum

et al. 2017

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Background Corynebacterium glutamicum is an important industrial workhorse and advanced genetic engineering tools are urgently demanded. Recently, the clustered regularly interspaced short palindromic repeats (CRISPR) and their CRISPR-associated proteins (Cas) have revolutionized the field of genome engineering. The CRISPR/Cas9 system that utilizes NGG as protospacer adjacent motif (PAM) and has good targeting specificity can be developed into a powerful tool for efficient and precise genome editing of C. glutamicum.ResultsHerein, we developed a versatile CRISPR/Cas9 genome editing toolbox for C. glutamicum. Cas9 and gRNA expression cassettes were reconstituted to combat Cas9 toxicity and facilitate effective termination of gRNA transcription. Co-transformation of Cas9 and gRNA expression plasmids was exploited to overcome high-frequency mutation of cas9, allowing not only highly efficient gene deletion and insertion with plasmid-borne editing templates (efficiencies up to 60.0 and 62.5%, respectively) but also simple and time-saving operation. Furthermore, CRISPR/Cas9-mediated ssDNA recombineering was developed to precisely introduce small modifications and single-nucleotide changes into the genome of C. glutamicum with efficiencies over 80.0%. Notably, double-locus editing was also achieved in C. glutamicum. This toolbox works well in several C. glutamicum strains including the widely-used strains ATCC 13032 and ATCC 13869.ConclusionsIn this study, we developed a CRISPR/Cas9 toolbox that could facilitate markerless gene deletion, gene insertion, precise base editing, and double-locus editing in C. glutamicum. The CRISPR/Cas9 toolbox holds promise for accelerating the engineering of C. glutamicum and advancing its application in the production of biochemicals and biofuels.Electronic supplementary materialThe online version of this article (10.1186/s12934-017-0815-5) contains supplementary material, which is available to authorized users.

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“…Products obtained with C. glutamicum have GRAS (generally regarded as safe) status and a long tradition in industry with well-established, large-scale cultivation processes. C. glutamicum has no significant extracellular proteolytic activity avoiding degradation of secreted protein or peptide products ( Becker and Wittmann, 2017 ; Lee and Kim, 2018 ). Moreover, the organism has been engineered for efficient growth and production on different renewable feedstocks and industrial waste streams including raw starches, pentoses from hemicellulosic wastes, mannitol from seaweed hydrolysates and amino sugars derived from chitin ( Hoffmann et al, 2021 ; Matano et al, 2014 ; Meiswinkel et al, 2013 ; Seibold et al, 2006 ).…”

Section: Introductionmentioning

confidence: 99%

Establishing recombinant production of pediocin PA-1 in Corynebacterium glutamicum

Goldbeck

Desef

Ovchinnikov

et al. 2021

Metabolic Engineering

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Bacteriocins are antimicrobial peptides produced by bacteria to inhibit competitors in their natural environments. Some of these peptides have emerged as commercial food preservatives and, due to the rapid increase in antibiotic resistant bacteria, are also discussed as interesting alternatives to antibiotics for therapeutic purposes. Currently, commercial bacteriocins are produced exclusively with natural producer organisms on complex substrates and are sold as semi-purified preparations or crude fermentates. To allow clinical application, efficacy of production and purity of the product need to be improved. This can be achieved by shifting production to recombinant microorganisms. Here, we identify Corynebacterium glutamicum as a suitable production host for the bacteriocin pediocin PA-1. C. glutamicum CR099 shows resistance to high concentrations of pediocin PA-1 and the bacteriocin was not inactivated when spiked into growing cultures of this bacterium. Recombinant C. glutamicum expressing a synthetic pedACD Cgl operon releases a compound that has potent antimicrobial activity against Listeria monocytogenes and Listeria innocua and matches size and mass:charge ratio of commercial pediocin PA-1. Fermentations in shake flasks and bioreactors suggest that low levels of dissolved oxygen are favorable for production of pediocin. Under these conditions, however, reduced activity of the TCA cycle resulted in decreased availability of the important pediocin precursor l -asparagine suggesting options for further improvement. Overall, we demonstrate that C. glutamicum is a suitable host for recombinant production of bacteriocins of the pediocin family.

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Industrial Microorganisms:Corynebacterium glutamicum

Cited by 25 publications

References 259 publications

The Mycobacterial Cell Envelope: A Relict From the Past or the Result of Recent Evolution?

The Mycobacterial Cell Envelope: A Relict From the Past or the Result of Recent Evolution?

Development of a CRISPR/Cas9 genome editing toolbox for Corynebacterium glutamicum

Establishing recombinant production of pediocin PA-1 in Corynebacterium glutamicum

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