Sequence Analysis of Lantibiotics: Chemical Derivatization Procedures Allow a Fast Access to Complete Edman Degradation

Meyer, Helmut E.; Heber, M.; Eisermann, B; Korte, H.; Metzger, Jörg W.; Jung, G.

doi:10.1006/abio.1994.1571

Cited by 77 publications

(76 citation statements)

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“…The easy loss of 146 amu upon acid treatment, which transformed 1 into a more hydrophobic species, together with acetonide formation observed with the intact molecule, suggested the presence of a sugar moiety. Lack of reactions with dithiothreitol and iodoacetamide 13 indicated the absence of disulfides and free cysteines, whereas the different reactivity observed with ethanethiol (EtSH) at basic 14 and neutral 15 pH indicated the presence of two thioethers. However, the many overlapping signals observed in the NMR spectrum of 1 prevented structure elucidation.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

A Glycosylated, Labionin-Containing Lanthipeptide with Marked Antinociceptive Activity

Iorio¹,

Sasso

Maffioli³

et al. 2013

ACS Chem. Biol.

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Among the growing family of ribosomally synthesized, post-translationally modified peptides, particularly intriguing are class III lanthipeptides containing the triamino acid labionin. In the course of a screening program aimed at finding bacterial cell wall inhibitors, we discovered a new lanthipeptide produced by an Actinoplanes sp. The molecule, designated NAI-112, consists of 22 amino acids and contains an N-terminal labionin and a C-terminal methyl-labionin. Unique among lanthipeptides, it carries a 6-deoxyhexose moiety N-linked to a tryptophan residue. Consistently, the corresponding gene cluster encodes, in addition to the LanKC enzyme characteristic of this lanthipeptide class, a glycosyl transferase. Despite possessing weak antibacterial activity, NAI-112 is effective in experimental models of nociceptive pain, reducing pain symptoms in mice in both the formalin and the chronic constriction injury tests. Thus, NAI-112 represents, after the labyrinthopeptins, the second example of a lanthipeptide effective against nociceptive pain.

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Section: ■ Results and Discussionmentioning

confidence: 99%

A Glycosylated, Labionin-Containing Lanthipeptide with Marked Antinociceptive Activity

Iorio¹,

Sasso

Maffioli³

et al. 2013

ACS Chem. Biol.

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“…The presence of such unusual residues may help explain why amino acid analyses of partially purified SLS preparations (2,28) have differed from the sequence predicted for the SagA propeptide (e.g., absence of cysteine). Previous attempts at sequencing SLS peptide by Edman degradations were unsuccessful (2), which may reflect cyclical thioether bridge structures or N-terminal blockage by a 2-oxobutyryl group as seen in other bacteriocins (35).…”

Section: Discussionmentioning

confidence: 99%

Genetic Locus for Streptolysin S Production by Group A Streptococcus

et al. 2000

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Group A streptococcus (GAS) is an important human pathogen that causes pharyngitis and invasive infections, including necrotizing fasciitis. Streptolysin S (SLS) is the cytolytic factor that creates the zone of betahemolysis surrounding GAS colonies grown on blood agar. We recently reported the discovery of a potential genetic determinant involved in SLS production, sagA, encoding a small peptide of 53 amino acids (S. D. Betschel, S. M. Borgia, N. L. Barg, D. E. Low, and J. C. De Azavedo, Infect. Immun. 66:1671-1679, 1998). Using transposon mutagenesis, chromosomal walking steps, and data from the GAS genome sequencing project (www .genome.ou.edu/strep.html), we have now identified a contiguous nine-gene locus (sagA to sagI) involved in SLS production. The sag locus is conserved among GAS strains regardless of M protein type. Targeted plasmid integrational mutagenesis of each gene in the sag operon resulted in an SLS-negative phenotype. Targeted integrations (i) upstream of the sagA promoter and (ii) downstream of a terminator sequence after sagI did not affect SLS production, establishing the functional boundaries of the operon. A rho-independent terminator sequence between sagA and sagB appears to regulate the amount of sagA transcript produced versus transcript for the entire operon. Reintroduction of the nine-gene sag locus on a plasmid vector restored SLS activity to the nonhemolytic sagA knockout mutant. Finally, heterologous expression of the intact sag operon conferred the SLS beta-hemolytic phenotype to the nonhemolytic Lactococcus lactis. We conclude that gene products of the GAS sag operon are both necessary and sufficient for SLS production. Sequence homologies of sag operon gene products suggest that SLS is related to the bacteriocin family of microbial toxins.Group A streptococcus (GAS), specifically Streptococcus pyogenes, is a common cause of pharyngitis, impetigo, and many other human respiratory tract and soft tissue infections. Recently, there has been a dramatic increase in reports of severe invasive GAS infections, including necrotizing fasciitis and toxic shock syndrome (44). Although GAS produces a wide array of virulence factors, those responsible for the rapid bacterial spread and tissue injury seen with invasive GAS infections are unknown.A hallmark feature of GAS in the clinical laboratory is the zone of beta-hemolysis observed surrounding colonies grown on the surface of blood agar media. The factor responsible for the beta-hemolytic phenotype is streptolysin S (SLS), the oxygen-stable cytolytic toxin of GAS. Despite detailed investigations over several decades, the exact chemical nature of SLS has remained a great mystery of GAS biology. SLS can exist in intracellular, cell-surface-bound, and extracellular forms (16), but attempts at purification are complicated by instability of the cytolytic activity in the absence of high-molecular-weight carriers, such as yeast RNA core or albumin (47).We have adopted a molecular genetic approach towards the identification of SLS and the study of...

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“…Samples for automated Edman degradation were applied to a TFA-treated glass fiber support (13). Prior to N-terminal sequencing, the peptide was derivatized by using the method of Meyer et al (23) as modified by Walk et al (42). Mass analyses and N-terminal sequencing were carried out at the Protein Microchemistry Facility (Department of Biochemistry, University of Otago).…”

Section: Methodsmentioning

confidence: 99%

Molecular and Genetic Characterization of a Novel Nisin Variant Produced by Streptococcus uberis

Wirawan

Klesse

Jack

et al. 2006

Appl Environ Microbiol

181

154

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Streptococcus uberis is one of the principal causative agents of bovine mastitis. In this study, we report that S. uberis strain 42 produces a lantibiotic, nisin U, which is 78% identical (82% similar) to nisin A from Lactococcus lactis. The 15.6-kb nisin U locus comprises 11 open reading frames, similar in putative functionality but differing in arrangement from that of the nisin A biosynthetic cluster. The nisin U producer strain exhibits specific resistance (immunity) to nisin U and cross-resistance to nisin A, a finding consistent with the 55% sequence similarity of their respective immunity peptides. Homologues of the nisin U structural gene were identified in several additional S. uberis strains, and in each case cross-protective immunity was expressed to nisin A and to the other producers of nisin U and its variants. To our knowledge, this is the first report both of characterization of a bacteriocin by S. uberis, as well as of a member of the nisin family of peptides in a species other than L. lactis.The lactic acid bacterium Streptococcus uberis, in nature primarily found on the lips and skin of cows, in raw milk, and on udder tissue (11), is also a major cause of bovine mastitis (4). Due to the ubiquity of this bacterium in the environment of the dairy cow, teat-end contamination poses a constant threat of infection, and the current level of disease caused by S. uberis remains a persistent problem impacting both on the economic production of milk and on the welfare of the dairy cow (21).Many lactic acid bacteria produce broad-spectrum proteinaceous antimicrobials called bacteriocins, some of which could provide valuable alternatives to traditional therapeutic antibiotics for the treatment of infectious diseases (30). Two such bacteriocins, nisin and lacticin 3147, which are produced by strains of Lactococcus lactis are potential candidates for mastitis control (5, 30). Nisin is the active ingredient in two commercial products: Consept (Applied Microbiology, Inc., New York, NY) and Wipe-Out (ImmuCell, Portland, OR). Lacticin 3147 has also been evaluated as a teat-seal formulation for the prevention of mastitis during the "dry" period in which the cow is not lactating (30).Nisin and lacticin 3147 both belong to the lantibiotic class of bacteriocins (10, 34). The lantibiotics are ribosomally synthesized, low-molecular-weight, heat-stable peptides characterized by their content of posttranslationally modified amino acids, including lanthionine and/or ␤-methyl-lanthionine (22,26,31). Nisin is the most intensively studied lantibiotic (20,35,37). Nisin Z (25) and nisin Q (45) are two natural variants of the original nisin A, differing in their propeptide components from nisin A by one and four amino acids, respectively. Lantibiotic loci typically comprise a structural gene (lanA) and other genes that encode proteins responsible for posttranslational modification of the prepeptide (lanB and lanC, or lanM), proteolytic processing (lanP), transport (lanT), producer selfprotection (lanI and lanEFG), and regulatio...

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Sequence Analysis of Lantibiotics: Chemical Derivatization Procedures Allow a Fast Access to Complete Edman Degradation

Cited by 77 publications

References 0 publications

A Glycosylated, Labionin-Containing Lanthipeptide with Marked Antinociceptive Activity

A Glycosylated, Labionin-Containing Lanthipeptide with Marked Antinociceptive Activity

Genetic Locus for Streptolysin S Production by Group A Streptococcus

Molecular and Genetic Characterization of a Novel Nisin Variant Produced by Streptococcus uberis

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