Mechanistic Studies of the Kinase Domains of Class IV Lanthipeptide Synthetases

Julian D. Hegemann, Liuqing Shi, Michael L. Gross, Wilfred Adrianus van der Donk

Research output: Contribution to journalArticle

Abstract

Lanthipeptides, which belong to the superfamily of ribosomally synthesized and posttranslationally modified peptides (RiPPs), are associated with various interesting biological activities. Lanthipeptides can be subdivided into four classes that are defined by the characteristics of the corresponding posttranslational modification enzymes. Class IV lanthipeptide synthetases consist of an N-terminal lyase, a central kinase, and a C-terminal cyclase domain. Here, we present the first in-depth characterization of such a kinase domain from the globisporin maturation enzyme SgbL that originates from Streptomyces globisporus sp. NRRL B-2293. Catalytic residues were identified by alignments with homologues and structural modeling. Their roles were confirmed by employing proteins with Ala substitutions in in vitro modification and fluorescence polarization binding assays. Furthermore, the protein region that is binding the leader peptide was identified by hydrogen-deuterium exchange-mass spectrometry experiments. By fusion of this protein region to the maltose binding protein, a protein was generated that can specifically bind the SgbA leader peptide, albeit with reduced binding affinity compared to that of full length SgbL. Combined, the results of this study provide a firmer grasp of how lanthipeptide biosynthesis is accomplished by class IV synthetases and suggest by homology analysis that biosynthetic mechanisms are similar in class III lanthipeptide processing enzymes.

Original languageEnglish (US)
Pages (from-to)1583-1592
Number of pages10
JournalACS chemical biology
Volume14
Issue number7
DOIs
StatePublished - Jun 5 2019

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Ligases
Phosphotransferases
Protein Sorting Signals
Proteins
Enzymes
Maltose-Binding Proteins
Lyases
Fluorescence Polarization
Deuterium
Biosynthesis
Streptomyces
Hand Strength
Post Translational Protein Processing
Bioactivity
Mass spectrometry
Hydrogen
Assays
Mass Spectrometry
Substitution reactions
Fusion reactions

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine

Cite this

Mechanistic Studies of the Kinase Domains of Class IV Lanthipeptide Synthetases. / Hegemann, Julian D.; Shi, Liuqing; Gross, Michael L.; van der Donk, Wilfred Adrianus.

In: ACS chemical biology, Vol. 14, No. 7, 05.06.2019, p. 1583-1592.

Research output: Contribution to journalArticle

Hegemann, Julian D. ; Shi, Liuqing ; Gross, Michael L. ; van der Donk, Wilfred Adrianus. / Mechanistic Studies of the Kinase Domains of Class IV Lanthipeptide Synthetases. In: ACS chemical biology. 2019 ; Vol. 14, No. 7. pp. 1583-1592.
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