Macrocyclization by asparaginyl endopeptidases

Abstract

Contents Summary923I.Introduction923II.Plant AEPs with macrocyclizing ability924III.Mechanism of macrocyclization by AEPs925IV.Conclusions927Acknowledgements927References927 Summary Plant asparaginyl endopeptidases (AEPs) are important for the post‐translational processing of seed storage proteins via cleavage of precursor proteins. Some AEPs also function as peptide bond‐makers during the biosynthesis of several unrelated classes of cyclic peptides, namely the kalata‐type cyclic peptides, PawS‐Derived Peptides and cyclic knottins. These three families of gene‐encoded peptides have different evolutionary origins, but all have recruited AEPs for their maturation. In the last few years, the field has advanced rapidly, with the biochemical characterization of three plant AEPs capable of peptide macrocyclization, and insights have been gained from the first AEP crystal structures, albeit mammalian ones. Although the biochemical studies have improved our understanding of the mechanism of action, the focus now is to understand what changes in AEP sequence and structure enable some plant AEPs to perform macrocyclization reactions.

Vacuolar processing enzymes in the plant life cycle

“…Their production requires a peptide ligation step in which peptide ligase(s) mediate the formation of a new peptide bond between the carboxyl and amino terminals within the precursor protein. VPEs in several plant species have a peptide ligation activity and are responsible for producing cyclic peptides (James et al , ). Jack bean β‐type VPE is required for post‐translational processing and peptide ligation of concanavalin A (Abe et al , ; Min & Jones, ; Takeda et al , ).…”

The macrocyclizing protease butelase 1 remains auto-catalytic and reveals the structural basis for ligase activity

“…Mutating a bulky Cys residue located in the substrate channel to a smaller Ala residue greatly increased the rates of macrocyclization activity of an AEP from Oldenlandia affinis (Yang et al, 2017). Furthermore, recent sequence analysis and molecular modeling has revealed a range of residues that together contribute to ligation efficiency (Jackson et al, 2018;James et al, 2018;Zauner et al, 2018a).…”

“…In addition to these cleavage functions, AEPs have attracted interest as enzymes that synthesize 43 cyclic peptides (reviewed by (James et al, 2018)). Several structural and molecular studies have 44 helped decipher the structural features necessary for macrocyclization.…”

“…Several structural and molecular studies have 44 helped decipher the structural features necessary for macrocyclization. We have recently 45 shown through structural and biochemical studies of a sunflower AEP that subtle amino acid 46 changes around the active site were able to alter the ratio of cyclized and cleaved reaction 47 products (Haywood et al, 2018) and that all activated AEPs are likely able to perform a 48 macrocyclization reaction at a favorable pH. Mutating a bulky Cys residue located in the 49 substrate channel to a smaller Ala residue greatly increased the rates of macrocyclization 50 transpeptidation and has been shown to circularize a number of non-native substrates of 66 various sequence compositions and sizes (Nguyen et al, 2014;Nguyen et al, 2015;Hemu et al, 67 2016).…”

Unravelling the mode of action of plant proteases

“…VPEs are also called asparaginyl endopeptidases (AEPs) and these VPEs/AEPs play an intriguing role in the creation of novel protein variants, often in seeds. In a Tansley insight article, James et al . (in this issue, pp.…”