Enzymatic and structural characterization of the major endopeptidase in the Venus flytrap digestion fluid

Michael W Risor, Line R Thomsen, Kristian W Sanggaard, Tania A Nielsen, Ida B Thøgersen, Marie V Lukassen, Litten Rossen, Irene Garcia-Ferrer, Tibisay Guevara, Carsten Scavenius, Ernst Meinjohanns, F Xavier Gomis-Ruth, Jan J Enghild

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

Abstract

Carnivorous plants primarily use aspartic proteases during digestion of captured prey. In contrast, the major endopeptidases in the digestive fluid of the Venus flytrap (Dionaea muscipula) are cysteine proteases (dionain-1 to -4). Here, we present the crystal structure of mature dionain-1 in covalent complex with inhibitor E-64 at 1.5 Å resolution. The enzyme exhibits an overall protein fold reminiscent of other plant cysteine proteases. The inactive glycosylated pro-form undergoes autoprocessing and self-activation, optimally at the physiologically relevant pH value of 3.6, at which the protective effect of the prodomain is lost. The mature enzyme was able to efficiently degrade a Drosophila fly protein extract at pH 4 showing high activity against the abundant Lys- and Arg-rich protein, myosin. The substrate specificity of dionain-1 was largely similar to that of papain with a preference for hydrophobic and aliphatic residues in subsite S 2 and for positively charged residues in S 1. A tentative structure of the pro-domain was obtained by homology modeling and suggested that a pro-peptide Lys residue intrudes into the S 2 pocket, which is more spacious than in papain. This study provides the first analysis of a cysteine protease from the digestive fluid of a carnivorous plant and confirms the close relationship between carnivorous action and plant defense mechanisms.

Original languageEnglish
JournalJournal of Biological Chemistry
Volume291
Issue5
Pages (from-to)2271-2287
Number of pages17
ISSN0021-9258
DOIs
Publication statusPublished - 1 Jan 2016

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