Fusaricidins from Paenibacillus polymyxa M‐1, a family of lipohexapeptides of unusual complexity—a mass spectrometric study

Abstract

Paenibacillus polymyxa are rhizobacteria with a high potential to produce natural compounds of biotechnological and medical interest. Main products of P. polymyxa are fusaricidins, a large family of antifungal lipopeptides with a 15-guanidino-3-hydroxypentadecanoic acid (GHPD) as fatty acid side chain. We use the P. polymyxa strain M-1 as a model organism for the exploration of the biosynthetic potential of these rhizobacteria. Using matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) about 40 new fusaricidins were detected which were fractionated by reversed-phase (rp) HPLC. Their structure was determined by MALDI-LIFT-TOF/TOF fragment analysis. The dominant fragment in the product ion spectra of fusaricidins appeared at m/z 256.3, 284.3 and 312.4, respectively, indicating variations in their fatty acid part. Two new subfamilies of fusaricidins were introduced which contain guanidino-3-hydroxyhepta- and nonadecanoic acid as fatty acid constituents. Apparently, the end-standing guanidine group is not modified as shown by direct infusion nano-electrospray ionization mass spectrometry (nano-ESI MS). The results of this study suggest that advanced mass spectrometry is the method of choice for investigating natural compounds of unusual diversity, like fusaricidins. Copyright © 2016 John Wiley & Sons, Ltd.

Genome Mining of the Lipopeptide Biosynthesis of Paenibacillus polymyxa E681 in Combination with Mass Spectrometry: Discovery of the Lipoheptapeptide Paenilipoheptin

“…The main products of P. polymyxa strains usually are the fusaricidins, a large family of lipohexapeptides of unusual complexity containing a guanidino C15, C17, or C19 β‐hydroxy fatty acid. They appear as a complex of more than 80 structural variants in culture filtrates of strain M1 . Fusaricidins are distinguished by potent activities against fungi of the genera Fusarium and Aspergillus as well as Gram‐positive bacteria.…”

“…Such compounds generally appear attached to the outer surfaceo fm icroorganisms. Therefore, they can efficiently be made available for testing from surface extracts,w hich are well suited for their rapid, sensitived etection by MALDI-TOF MS. [17,18] Mass spectrometry structure analysiss tarted with the determinationo ft heir amino-acid components by their immoniumi ons, followed by the detectiono fn earest-neighbor relationshipsd erived from the fragment spectra.T his information formed the basis for de novo sequencing of the peptidep roducts for their identification and structuralc haracterization.…”

“…The module arrangementa long the assembly lines in Figure 1r eflects the sequence of the peptidep roducts. In this way,n onribosomal peptides ynthetase (NRPS) cluster A( 81155-111 975) and D (4 536 009-4 562 187) were attributed to the biosynthesis of the well-known fusaricidins [17,18,[22][23][24][25][26] and polymyxins, [19,27,28] which have been investigated in detail in previousw ork. Fusaricidin is produced by as ingle six-module enzyme, whereas in the biosynthesis of polymyxin, three multifunctional enzymes cooperate.…”

“…In accordance with the genome mining studies, four product families were detected in the extracts (Figure 2A-D). They were presented with increasing mass numbers, as they appear in the mass spectrum.T he ensembles found in the mass ranges m/z 860-1050 ( Figure 2A,c orresponding to gene cluster A), m/z 1140-1200 ( Figure 2B,c orresponding to gene cluster D), and m/z 1580-1680 ( Figure 2D,c orrespondingt og ene cluster B) were attributedt ot he well-known fusaricidins, [17,18,[22][23][24][25][26] polymyxins, [19,27,28] and tridecaptins. [12,[34][35][36][37] The main fusaricidin specieso fP.…”

“…polymyxa E681 are fusaricidins A, B, and Ei nc ontrastt os train M1, which exhibits fusaricidins A, B, C, and Da st he dominant products. [17,18] The polymyxins produced by strain E681 are of type A. They appear only in low amount.T heir mass signals were strongly superposed by those of yet-unknown compounds showing mass signals between m/z 1100 and 1180 ( Figure 2C).…”