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Metacaspases of the Fungus S. commune Are Activated by Calcium and Specific for Arginine
Author(s) -
Leang Lakhena,
Fox Kristin M.
Publication year - 2018
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2018.32.1_supplement.526.44
Subject(s) - biochemistry , arginine , chemistry , enzyme , cysteine , proteases , saccharomyces cerevisiae , cysteine protease , schizophyllum commune , histidine , amino acid , gene
Metacaspases belong to a class of cysteine peptidase protein enzymes and are known to be involved in several programmed cell death (PCD) mechanisms in plants and lower eukaryotes. Similar to caspases, metacaspases contain a cysteine‐histidine catalytic dyad and can undergo autoprocessing for increased activity. However, these proteases cleave after arginine or lysine residues, while caspases are aspartate specific. Previous studies have shown that Ca 2+ plays a critical role in metacaspase activity. The metacaspase structures of MCA2 ( Trypanosoma brucei) and YCA1 ( Saccharomyces cerevisiae ) have been found to contain calcium‐binding site(s) necessary for complete enzyme activity. A subset of metacaspases possesses an N‐terminal prodomain containing a proline rich sequence. We have identified five metacaspase genes in this subset from the split‐gilled fungus Schizophyllum commune (ScMC1‐5). All genes for ScMC1‐5 have been cloned with and without the prodomain (Δpro) into a SUMO vector construct, expressed, purified and tested for enzymatic activity. The goal of this project was to characterize ScMC1‐5Δpro by determining substrate specificity, optimal Ca 2+ concentrations, and optimal pH levels for each enzyme using a metacaspase fluorogenic activity assay. Results show that all ScMC1‐5Δpro showed arginine and lysine specificity, with higher efficiency for arginine. All enzymes performed best within a pH range of 7 and 8. Optimal calcium concentrations ranged from 5 mM to 50 mM Ca 2+ . Only ScMC1Δpro is completely inactive at 0 mM calcium. ScMC1&5Δpro have a much higher optimal concentration (50 mM), compared to ScMC2, 3 and 4Δpro, suggesting that calcium may have different binding affinities and play different roles in each enzyme. These data agree with current literature for other similarly characterized metacaspases. Increasing our understanding of metacaspase enzymes could provide vital information for the development of future anti‐fungal and antibacterial treatments. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .