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The isolation of the cyclic lipopeptide arthrofactin from the bacterium Arthrobacter sp. MIS38 was first described in 1993 in the context of its high surface activity. (Morikawa, 1993) Later, the genus of the producer strain was revised to be Pseudomonas fluorescens MIS38. (Morikawa, 2000) The structure of the lipopeptide was revised in 2012, based on NMR spectroscopy and chiral HPLC analyses. (Lange, 2012)

Original publicationMorikawa, 1993
Original sourcePseudomonas fluorescens MIS38
Sequence determined byMS-MS spectrometry
Stereochemistry determined byGC-MS
Other known sources (non-putative)n.a.
NMR data availableDMSO-d6 (Lange, 2012)

Chemical properties

Molecular formulaC66H111N11O20
Mono-isotopic mass1353.8007 (isobaric to other CLiPs, see text)
Molecular weight1354.6 g mol-1
SolubilitySoluble in MeOH, DMSO, DMF, buffer.
CMC1.0 .10-5 M



Arthrofactin is a cyclic lipodepsipeptide (CLiP) produced by Pseudomonas sp. MIS38,. It consists of a undecapeptide sequence, linked to at a 3R-hydroxydecanoic acid (3-OH C10:0) moiety at the N-terminus, as determined by MS-MS spectrometry. Originally, it was believed that the C-terminal carbonyl forms an ester bond with the hydroxyl moiety of the fatty acid tail, as observed for some Bacillus CLiPs (surfactins). However, this was revised in 2012, where it was demonstrated that the peptide features an ester bond between the C-terminal carbonyl and the side chain of threonine at position 3, forming a macrocycle containing 9 of the 11 amino acid. The primary structure of arthrofactin is thus 3R-hydroxydecanoyl – D-Leu1- D-Asp2 – D-aThr3 – D-Leu4 – D-Leu5 – D-Ser6 – L-Leu7 – D-Ser8 – L-Ile9 – L-Ile10 – L-Asp11.

In addition to the major product arthrofactin A, the producer strain is also known to biosynthesize several derivatives as side products of the same gene cluster .This phenomenon is commonly observed in CLiP biosynthesis and can be explained by the relaxed substrate specificity of the C1 domain of module 1 and A domains in each module, which lead to altered fatty acid moieties and different amino acid compositions of the CLiP, respectively. Using a combination of chiral HPLC and MS/ MS techniques, it was determined that arthrofactin B differs by the replacement of L-Asp11 with L-Glu. Arthrofactin C and D both differ from the main compound in their fatty acid moiety, featuring a 3-OH C12:1 and 3-OH C12:0 fatty acid tail, respectively, instead of 3-OH C10:0.

Chemical structure of arthrofactin A
Schematic sequences of arthrofactin A and homologues, where shapes indicate amino acid configuration (circles = L-AA, squares = D-AA) and colors indicate amino acid polarity (green = hydrophobic, red = polar)

The arthrofactins are members of the amphisin group, a collection of structurally similar lipopeptides including amphisin, tensin, lokisin, anikasin, milkisins and stechlisins. The group is also reported to contain two ill-characterized CLiPs: hordersin and pholipeptin. Within the amphisin group, variations mostly occur at position 8 (Gln vs. Ser), position 9 (Leu vs Ile) and position 11 (Asp vs Glu). Arthrofactin A is isobaric to lokisin, anikasin and pholipeptin. Consequently, these cannot be discriminated from one another using mass spectrometry alone.


No information is available on the antagonistic activities of arthrofactin.

Physicochemical properties

Arthrofactin is an exceptionally effective cyclic lipodepsipeptide biosurfactant. It reduces the surface tension of water from 72 to 24 mN/m at a critical micelle concentration of 1×10-5 M. Additionally, it displays a high oil displacement. (Morikawa, 1993)


The biosynthesis of arthrofactin has been studied extensively. (Roongsawang, 2003, Balibar, 2005, Roongsawang, 2007, Lange, 2012) The lipopeptide is produced by non-ribosomal peptide synthetase, multimodular peptides responsible for the biosynthesis of certain secondary metabolite. The arthrofactin synthetase genes contain three biosynthetic gene clusters (arfA, arfB, and arfC) that encode for two, four and five functional NRPS modules, respectively. Every module contains different domains, each with a specific function. The adenylation (A) domain is responsible for amino acid recognition and adenylation at the expense of adenosine triphosphate (ATP) to form an acyl-adenylate intermediate. Then, the adenylated amino acid covalently binds to the adjacent thiolation (T) domain. Peptide bond formation of two consecutively bound amino acids is catalysed by the condensation (C) domain. Lastly, cyclization and release of the product peptide are performed by C-terminal thioesterase (Te) domain. None of the 11 modules of the arthrofactin NRPS contain epimerisation domains, typically responsible for the conversion of L-amino acids to their D-configuration. Instead, the D-configuration of the amino acids in arthrofactin is generated by specific C domains that have dual catalytic activities, i.e., both condensation and epimerization. These C/E domains are involved in the epimerization of the amino acid that is loaded onto the T domain of the preceding module. In case of arthrofactin, two Te domains are tandemly located at the C-terminal end of ArfC, responsible for the cyclisation and release of the peptide product. The first Te-domain (Te1) was shown to be essential for the completion of macrocyclization and the release of the final product. The second Te-domain (Te2) was suggested to be evolved to improve the macrocyclization efficiency.

Finally, the synthesis of the fatty acid chain is believed to be part of the primary metabolism since no genes responsible for the biosynthesis of the fatty acid moiety have been found in the vicinity of the NRPS genes of Pseudomonas bacteria.

NMR fingerprint data

Recently, it was established that the planar structure and stereochemistry of CLiPs can be assessed by simple comparison to a reference. (De Roo, 2022) More specifically, by matching NMR spectra of a CLiP from a newly isolated bacterial source with those of existing (reference) CLiPs, one can determine whether they are identical or not. A detailed explanation on what NMR fingerprint matching is, and how to use it, can be found here.

Below, we provide the reference NMR data of arthrofactin. This data is recorded in DMSO-d6 at room temperature, and can be used to asses similarities of newly isolated CLiPs to arthrofactin.

ATTENTION: The solvent of choice for NMR reference spectra is DMF-d7. However, since no spectra are available in this solvent, we provide the NMR data as recorded in DMSO-d6. If you wish to compare spectra of your own CLiP to the data of arthrofactin A, please use identical conditions! Moreover, since the arthrofactin homologues differ only in the identity of the fatty acid moiety, the chemical shifts of arthrofactins B – D are identical to those of arthrofactin A. Be sure to check the length and saturation of the fatty acid tail via a complementary technique such as mass spectrometry.


Balibar, et al. “Generation of D amino acid residues in assembly of arthrofactin by dual condensation/epimerization domains.” Chemistry & Biology12, 11 (2005): https://dx.doi.org/10.1016/j.chembiol.2005.08.010.

Lange, et al. “Predicting the structure of cyclic lipopeptides by bioinformatics: structure revision of arthrofactin.” ChemBioChem13, 18 (2012): https://dx.doi.org/10.1002/cbic.201200532.

Morikawa, et al. “A new lipopeptide biosurfactant product by Arthrobacter  sp. strain MIS38.” Journal of Bacteriology175, 20 (1993).

Morikawa, et al. “A study on the structure-function relationship of lipopeptide biosurfactants.” Biochimica et Biophysica Acta1488 (2000).

Roongsawang, et al. “Cloning and characterization of the gene cluster encoding arthrofactin synthetase from Pseudomonas sp. MIS38.” Chemistry & Biology10, 9 (2003): https://dx.doi.org/10.1016/j.chembiol.2003.09.004.

Roongsawang, et al. “In vivo characterization of tandem C-terminal thioesterase domains in arthrofactin synthetase.” ChemBioChem8, 5 (2007): https://dx.doi.org/10.1002/cbic.200600465.

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