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Tolaasin A-E
General info
Chemical properties
| CAS | n.a. |
| Molecular formula | Tolaasin A: C91H155N21O26 Tolaasin B: C93H163N21O25 Tolaasin C: C94H165N21O26 Tolaasin D: C94H163N21O25 Tolaasin E: C92H159N21O24 |
| Molecular weight | Tolaasin A: 1959.32 g/mol Tolaasin B: 1975.41 g/mol Tolaasin C: 2005.43 g/mol Tolaasin D: 1987.42 g/mol Tolaasin E: 1943.37 g/mol |
| Mono-isotopic mass | Tolaasin A: 1958.1452 Da Tolaasin B: 1974.2128 Da Tolaasin C: 2004.2234 Da Tolaasin D: 1986.2128 Da Tolaasin E: 1942.1867 Da |
| Solubility | Water, Methanol, Acetonitrile, DMSO, DMF,TFE |
| CMC | n.a. |
| Minimal surface tension | n.a. |
| 3D conformation | n.a. |
| NMR data available in literature | DMSO (Bassarello, 2004) |
Introduction
Tolaasins are cyclic lipodepsipeptides (CLiP) produced by Pseudomonas tolaasii and are the causal agents for brown blotch disease in mushrooms. (Wong, 1979) These CLiPs features 18 amino acids, whereby the N-terminus is capped by a (3R)-hydroxy fatty acid moiety. The C terminus forms a macrocycle by formation of an ester bond to a side chain hydroxyl function. The main CLiP produced by Pseudomonas tolaasii is tolaasin I. Besides this CLiP, several minor compounds are also produced. These compounds are called tolaasins A-E and represent structural homologues that typically result by flexibility in the A or Cstart domains of the non-ribosomal peptide synthetases.
Tolaasins A-E belong to the Tolaasin (18:5) group that consists of tolaasin I-II (Nutkins, 1991), tolaasin A-E (Bassarello, 2004), tolaasin F (Scherlach, 2013) and sessilin (D’Aes, 2014).
Chemical structure
The structures of tolaasins A-E were elucidated in 2004 by means of mass spectrometry, amino acid sequencing and 1H NMR spectroscopy. Tolaasins are relatively large CLiP, containing 18 amino acids, 5 of which form a lactone macrocycle. The peptide cyclization involves an ester bond between the C-terminus and the alcohol side-chain function of D-aThr14. Unnatural amino acids, such as homoserine (Hse), 2,4-diaminobutyric acid (Dab) and 2,3-didehydroaminobutyric acid (Dhb) are common in the lipopeptides of the Tolaasin group, while they do not occur in e.g. the Viscosin, Orfamide or Amphisin group, indicating a more complex assembly process. The N-terminus of the tolaasins are typically capped by a 3R-hydroxy octanoic acid (3R-OH C8:0).
Pseudomonas tolaasii produces tolaasin I as main CLiP. In addition, it produces a number of minor compounds, called tolaasin A-E. Tolaasin A is a minor structural homologue that features an N-terminal 5-pentadioic acid (adipic acid; 5-COOH C5:0) instead of an 3-hydroxy octanoic acid (3-OH C8:0). Tolaasin B and D are tolaasin I homologues whereby Ile15 is replaced by Val15 or Ile15, respectively. Tolaasin E features two structural modifications in comparison to the major tolaasin I, whereby both Ile15 and Hse16 are replaced by Leu15 and Gly16. Finally, tolaasin C is a linear homologue of tolaasin I, whereby the ester bond is not formed.
At physiological pH, tolaasin A and C feature a single negative charge in addition to the two positive charges present in the other tolaasin structures.
Biological activity
More than 100 years ago, the Gram-negative Pseudomonas tolaasii Paine bacterium was revealed as being the organism responsible for causing brown-blotch disease on cultivated mushrooms, effectively making these unappealing and ruining their consumption. Since then, its detrimental effect on agricultural crops has also been reported for other crops, including strawberries, cauliflower and tobacco. The bacterium is endemic to the compost ground used for cultivation where it is non-pathogenic. Under the influence of multifactorial conditions where temperature and relative humidity are known to play a decisive role, P. tolaasii switches to a pathogenic state, where it secretes several virulence factors, i.e. secondary metabolites that effect the colonisation and infection at the molecular level. Tolaasin, an 18 residue cyclic lipopeptide, is the main virulence factor of P. tolaasii. Upon secretion by the bacterium, it damages the mushroom cellular membranes, thereby initiating a series of intracellular events that are well documented and ultimately lead to formation of pits in the mushroom caps with brown coloured discoloration resulting from the production of melanin. (Soler-Rivas, 1999) In addition to antifungal activity, antagonistic activity against both Gram-positive and Gram-negative bacteria has been described. (Geudens, 2018)
The results of the antimicrobial activity of tolaasins A-E suggest the importance of the lactone and the N-terminus acyl moiety. Tolaasin A, which features a pentadioic acid instead of β-hydroxyoctanoic acid, showed a reduced activity with respect to tolaasin I. Moreover, tolaasin C, the linear analogue of tolaasin I, was found to be biologically inactive. In Nature, the effect of the lactone ring opening on the activity of tolaasin has been associated with the detoxification mechanism of competing microorganisms. (Tomita, 2018, Hermenau, 2020)
References
Bassarello, et al. “Tolaasins A-E, five new lipodepsipeptides produced by Pseudomonas tolaasii.” Journal of Natural Products67, 5 (2004).
D’Aes, et al. “To settle or to move? The interplay between two classes of cyclic lipopeptides in the biocontrol strain Pseudomonas CMR12a.” Environmental Microbiology16, 7 (2014): https://dx.doi.org/10.1111/1462-2920.12462.
Geudens, et al. “Cyclic lipodepsipeptides from Pseudomonas spp. – Biological Swiss-Army Knives.” Frontiers in Microbiology9, 1867 (2018): https://dx.doi.org/10.3389/fmicb.2018.01867.
Hermenau, et al. “Helper bacteria halt and disarm mushroom pathogens by linearizing structurally diverse cyclolipopeptides.” Proceedings of the National Academy of Sciences of the United States of America117, 38 (2020): https://dx.doi.org/10.1073/pnas.2006109117.
Nutkins, et al. “Structure determination of tolaasin, an extracellular lipodepsipeptide produced by the mushroom pathogen, Pseudomonas tolaasii Paine.” Journal of the American Chemical Society113, 7 (1991).
Scherlach, et al. “Biosynthesis and mass spectrometric imaging of tolaasin, the virulence factor of brown blotch mushroom disease.” ChemBioChem14, 18 (2013): https://dx.doi.org/10.1002/cbic.201300553.
Soler-Rivas, et al. “Biochemical and physiological aspects of brown blotch disease of Agaricus bisporus.” FEMS Microbiology Reviews23, 5 (1999): https://dx.doi.org/10.1111/j.1574-6976.1999.tb00415.x.
Tomita, et al. “Detoxification process of tolaasins, lipodepsipeptides, by Microbacterium sp. K3-5.” Bioscience, Biotechnology and Biochemistry82, 8 (2018): https://dx.doi.org/10.1080/09168451.2018.1460575.
Wong, et al. “Identification of Pseudomonas tolaasi: the white line in agar and mushroom tissue block rapid pitting tests.” Journal of Applied Bacteriology47 (1979).