title: Isolation, purification and structure of exochelin MS, the extracellular siderophore from Mycobacterium smegmatis.
abstract: The extracellular siderophore from Mycobacterium smegmatis, exochelin MS, was isolated from iron-deficiently grown cultures and purified to > 98% by a combination of ion-exchange chromatography and h.p.l.c. The material is unextractable into organic solvents, is basic (pI = 9.3-9.5), has a lambda max at 420 nm and a probable Ks for Fe3+ of between 10(25) and 10(30). Its structure has been determined by examination of desferri- and ferri-exochelin and its gallium complex. The methods used were electrospray-m.s. and one- and two-dimensional (NOESY, DQF-COSY and TOCSY) 1H n.m.r. The constituent amino acids were examined by chiral g.l.c analysis of N-trifluoroacetyl isopropyl and N-pentafluoropropionyl methyl esters after hydrolysis, and reductive HI hydrolysis, of the siderophore. The exochelin is a formylated pentapeptide: N-(delta-N-formyl,delta N-hydroxy-R-ornithyl) -beta-alaninyl-delta N-hydroxy-R-ornithinyl-R-allo-threoninyl-delta N-hydroxy-S-ornithine. The linkages involving the three ornithine residues are via their delta N(OH) and alpha-CO groups leaving three free alpha-NH2 groups. Although there are two peptide bonds, these involve the three R (D)-amino acids. Thus the molecule has no conventional peptide bond, and this suggests that it will be resistant to peptidase hydrolysis. The co-ordination centre with Fe3+ is hexadenate in an octahedral structure involving the three hydroxamic acid groups. Molecular modelling shows it to have similar features to other ferric trihydroxamate siderophores whose three-dimensional structures have been established. The molecule is shown to have little flexibility around the iron chelation centre, although the terminal (Orn-3) residue, which is not involved in iron binding except at its delta N atom, has more motional freedom.
authors: Department of Chemistry, Cambridge University, U.K.
authors: G J
authors: GJ
authors: Sharman
authors: D H
authors: DH
authors: Williams
authors: D F
authors: DF
authors: Ewing
authors: C
authors: C
authors: Ratledge
mesh_terms: D000595:Amino Acid Sequence; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009161:Mycobacterium / Q000737:chemistry*; D010456:Peptides, Cyclic / Q000737:chemistry* / Q000302:isolation & purification*; D011487:Protein Conformation; D017262:Siderophores / Q000737:chemistry* / Q000302:isolation & purification*; D013056:Spectrophotometry, Ultraviolet; D013237:Stereoisomerism
chemical_list: D010456:Peptides, Cyclic; D017262:Siderophores; C010953:exochelins