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Involvement of a Novel Class C Beta-Lactamase in the Transglutaminase Mediated Cross-Linking Cascade of Streptomyces mobaraensis DSM 40847.

Zindel, Stephan and Ehret, Vera and Ehret, Marina and Hentschel, Madeleine and Witt, Samantha and Krämer, Andreas and Fiebig, David and Jüttner, Norbert and Fröls, Sabrina and Pfeifer, Felicitas and Fuchsbauer, Hans-Lothar (2016):
Involvement of a Novel Class C Beta-Lactamase in the Transglutaminase Mediated Cross-Linking Cascade of Streptomyces mobaraensis DSM 40847.
In: PloS one, pp. e0149145, 11, (2), ISSN 1932-6203, [Article]

Abstract

Streptomyces mobaraensis DSM 40847 secretes transglutaminase that cross-links proteins via γ-glutamyl-ε-lysine isopeptide bonds. Characterized substrates are inhibitory proteins acting against various serine, cysteine and metalloproteases. In the present study, the bacterial secretome was examined to uncover additional transglutaminase substrates. Fractional ethanol precipitation of the exported proteins at various times of culture growth, electrophoresis of the precipitated proteins, and sequencing of a 39 kDa protein by mass spectrometry revealed the novel beta-lactamase Sml-1. As indicated by biotinylated probes, Sml-1, produced in E. coli, exhibits glutamine and lysine residues accessible for transglutaminase. The chromogenic cephalosporin analogue, nitrocefin, was hydrolyzed by Sml-1 with low velocity. The obtained Km and kcat values of the recombinant enzyme were 94.3±1.8 μM and 0.39±0.03 s-1, respectively. Penicillin G and ampicillin proved to be weak inhibitors of nitrocefin hydrolysis (Ki of 0.1 mM and 0.18 mM). Negligible influence of metals on β-lactamase activity ruled out that Sml-1 is a Zn2+-dependent class B beta-lactamase. Rather, sequence motifs such as SITK, YSN, and HDG forming the active core in a hypothetical structure may be typical for class C beta-lactamases. Based on the results, we assume that the novel transglutaminase substrate ensures undisturbed growth of aerial hyphae in Streptomyces mobaraensis by trapping and inactivating hostile beta-lactam antibiotics.

Item Type: Article
Erschienen: 2016
Creators: Zindel, Stephan and Ehret, Vera and Ehret, Marina and Hentschel, Madeleine and Witt, Samantha and Krämer, Andreas and Fiebig, David and Jüttner, Norbert and Fröls, Sabrina and Pfeifer, Felicitas and Fuchsbauer, Hans-Lothar
Title: Involvement of a Novel Class C Beta-Lactamase in the Transglutaminase Mediated Cross-Linking Cascade of Streptomyces mobaraensis DSM 40847.
Language: English
Abstract:

Streptomyces mobaraensis DSM 40847 secretes transglutaminase that cross-links proteins via γ-glutamyl-ε-lysine isopeptide bonds. Characterized substrates are inhibitory proteins acting against various serine, cysteine and metalloproteases. In the present study, the bacterial secretome was examined to uncover additional transglutaminase substrates. Fractional ethanol precipitation of the exported proteins at various times of culture growth, electrophoresis of the precipitated proteins, and sequencing of a 39 kDa protein by mass spectrometry revealed the novel beta-lactamase Sml-1. As indicated by biotinylated probes, Sml-1, produced in E. coli, exhibits glutamine and lysine residues accessible for transglutaminase. The chromogenic cephalosporin analogue, nitrocefin, was hydrolyzed by Sml-1 with low velocity. The obtained Km and kcat values of the recombinant enzyme were 94.3±1.8 μM and 0.39±0.03 s-1, respectively. Penicillin G and ampicillin proved to be weak inhibitors of nitrocefin hydrolysis (Ki of 0.1 mM and 0.18 mM). Negligible influence of metals on β-lactamase activity ruled out that Sml-1 is a Zn2+-dependent class B beta-lactamase. Rather, sequence motifs such as SITK, YSN, and HDG forming the active core in a hypothetical structure may be typical for class C beta-lactamases. Based on the results, we assume that the novel transglutaminase substrate ensures undisturbed growth of aerial hyphae in Streptomyces mobaraensis by trapping and inactivating hostile beta-lactam antibiotics.

Journal or Publication Title: PloS one
Volume: 11
Number: 2
Divisions: 10 Department of Biology
10 Department of Biology > Microbiology and Archaea
Date Deposited: 23 Feb 2016 10:36
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