Elucidating the specificity determinants of the AtxE2 lasso peptide isopeptidase

Mikhail O. Maksimov, Joseph D. Koos, Chuhan Zong, Bozhena Lisko, A. James Link

Research output: Contribution to journalArticlepeer-review

24 Scopus citations


Lasso peptide isopeptidase is an enzyme that specifically hydrolyzes the isopeptide bond of lasso peptides, rendering these peptides linear. To carry out a detailed structure-activity analysis of the lasso peptide isopeptidase AtxE2 from Asticcacaulis excentricus, we solved NMR structures of its substrates astexin-2 and astexin-3. Using in vitro enzyme assays, we show that the C-terminal tail portion of these peptides is dispensable with regards to isopeptidase activity. A collection of astexin-2 and astexin-3 variants with alanine substitutions at each position within the ring and the loop was constructed, and we showed that all of these peptides except for one were cleaved by the isopeptidase. Thus, much like the lasso peptide biosynthetic enzymes, lasso peptide isopeptidase has broad substrate specificity. Quantitative analysis of the cleavage reactions indicated that alanine substitutions in loop positions of these peptides led to reduced cleavage, suggesting that the loop is serving as a recognition element for the isopeptidase.

Original languageEnglish (US)
Pages (from-to)30806-30812
Number of pages7
JournalJournal of Biological Chemistry
Issue number52
StatePublished - Dec 25 2015

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Biochemistry
  • Cell Biology


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