Characterization of exochelins of Mycobacterium avium: evidence for saturated and unsaturated and for acid and ester forms

Abstract

Mycobacterium avium secretes iron-binding siderophores called exochelins. The exochelins from M. avium have previously been reported to have unsaturated side chains that terminate in carboxylic acid. In contrast, our data show the side chains to be both saturated and unsaturated and to terminate with either a carboxylate or methyl ester.Mycobacterium avium is a slowly growing mycobacterium and an important human pathogen. Because M. avium is one of the most common opportunistic pathogens associated with AIDS, the prevalence of M. avium infection has skyrocketed in recent years (5).Like many pathogens, M. avium produces high-affinity ironbinding molecules known as siderophores to help it acquire iron in the host (9). Free iron is extremely limited in the host because of the high affinity with which it complexes to host iron-binding proteins. Other investigators (2,7,8,10) have shown that mycobacteria produce small water-soluble siderophores called exochelins. They have proposed that exochelins bind iron in the extracellular environment and deliver the iron to mycobactin, another high-affinity iron-binding molecule, which is located in the cell wall (or envelope) of the mycobacterium (8). Gobin and Horwitz have recently demonstrated that the exochelins of Mycobacterium tuberculosis remove iron from human transferrin and lactoferrin and donate iron to mycobactins in living intact organisms (3).Exochelins can be classified into two general categories depending on their extractability into organic solvents (11). Saprophytic mycobacteria produce exochelins which cannot be extracted into any organic solvent. In contrast, slowly growing, pathogenic mycobacteria, like M. tuberculosis and M. avium, produce exochelins that are extractable into chloroform (2). Mycobactins have been extensively studied (14), but because of purification problems, physiological and structural studies of exochelins have only recently been performed. The structures of chloroform-insoluble exochelins from the nonpathogens Mycobacterium smegmatis (13) and Mycobacterium neoaurum (12) have recently been described. These structures differ greatly from those of the chloroform-soluble exochelins isolated from the pathogens M. tuberculosis (4) and M. avium (6). More precisely, the exochelins of the nonpathogens are peptides, whereas the exochelins from the pathogenic mycobacteria resemble mycobactins and contain both amino acid and non-amino acid moieties. Although both our group and Lane et al. (6) found that the core structure of the exochelins of the mycobacteria resembles that of the mycobactins, the results of our two groups differ with respect to the R 1 alkyl side chain, which critically influences the water solubility of exochelins and allows them to function in the extracellular environment. Whereas we previously had reported that the R 1 alkyl side chain of the exochelins of M. tuberculosis exists in both saturated and unsaturated forms that terminate predominantly with a methyl ester but additionally with a free carboxylic acid, Lane et al. (...

Collision-activated dissociation, infrared multiphoton dissociation, and electron capture dissociation of the Bacillus anthracis siderophore petrobactin and its metal ion complexes

“…The same research group also showed that coupling of high-performance liquid chromatography (HPLC) with ESI MS is an effective strategy for separation and identification of siderophores in complex samples [19]. Gibson and coworkers applied MS/MS to characterize exochelin siderophores and found that its side chains are both saturated and unsaturated and terminated with either a caboxylate group or a methyl ester [11,21]. These results contrasted with the previous observation that exochelins are unsaturated and terminated with carboxylic acid [16].…”

“…Electrospray ionization mass spectrometry (ESI MS) and tandem mass spectrometry (MS/MS) have proved to be powerful techniques for detecting and identifying siderophores [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23]. For example, Gledhill demonstrated the utility of these techniques for characterizing hydroxamate-type siderophores, including ferrioxamine, ferrichrome, and iron(III) rhodotoluate [9].…”

Discovery and characterization of a unique mycobacterial heme acquisition system

“…Iron, essential for pathogen survival, is sequestered from its host by well-characterized siderophore-mediated iron acquisition pathways. Pathogenic mycobacteria including Mtb synthesize two types of siderophores-the cell-bound water-insoluble mycobactins and the secreted water-soluble exochelins, which have a mycobactin-like backbone (1,2), and are referred to as exomycobactins hereafter. Exomycobactins, the most abundant molecules secreted by Mtb on a molar basis, can remove iron from human transferrin and lactoferrin and transport it to either mycobactins in the Mtb cell wall (3) or the iron transport system, IrtA/B (4).…”

Identification and Characterization of a Major Cell Wall-Associated Iron-Regulated Envelope Protein (Irep-28) in Mycobacterium tuberculosis

“…Calder and Horwitz showed the iron-regulated expression of Irp10 and Mta72 in Mycobacterium tuberculosis by single-dimension electrophoresis (3) and later, using two-dimensional gel electrophoresis combined with mass spectrometry and sequence information (27), demonstrated the upregulation of a putative cation transporting ATPase, a mycobacterial homologue of phosphoenolpyruvate carboxykinase, and an NADPdependent dehydrogenase in bacteria grown in low-iron medium, whereas these authors observed increased levels of FurA, a homologue of EF-Tu, and an aconitase in iron-rich medium.…”