title: Biochemical evidence of key residues for the activation and autoprocessing of tomato type II metacaspase.
abstract: To investigate the autolysis pattern and activation of metacaspase in higher plants, the biochemical characteristics of purified recombinant type II metacaspase (LeMCA1) from tomato were explored. Western blotting analysis indicated that four cleaved bands were formed; two N-terminal fragments and two C-terminal fragments. N-terminal sequencing confirmed that LeMCA1 cleaves at Lys223 and Arg332. Site mutants indicated that catalytic Cys139, cleaved Lys223, Arg332 and predicted calcium binding Asp116/Asp117 are the key residues that are responsible for its Ca²⁺ and pH dependent activation. The cleavage of the full-size fragment seemed crucial for the activation of LeMCA1 in vitro.
authors.affiliation: College of Food Science and Nutritional Engineering, China Agricultural University, No. 17 Qinghua East Road, Beijing 100083, China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Wen
authors.forename: Qiu-Min
authors.initials: QM
authors.lastname: Ma
authors.forename: Ya-Li
authors.initials: YL
authors.lastname: Zhang
authors.forename: Ji-Ping
authors.initials: JP
authors.lastname: Yang
authors.forename: Guang-Hua
authors.initials: GH
authors.lastname: Zhao
authors.forename: Da-Qi
authors.initials: DQ
authors.lastname: Fu
authors.forename: Yun-Bo
authors.initials: YB
authors.lastname: Luo
authors.forename: Gui-Qin
authors.initials: GQ
authors.lastname: Qu
mesh_terms: D002118:Calcium / Q000737:chemistry; D020169:Caspases / Q000737:chemistry* / Q000235:genetics; D004789:Enzyme Activation; D018551:Solanum lycopersicum / Q000201:enzymology* / Q000235:genetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010455:Peptides / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry
chemical_list: D010455:Peptides; D010940:Plant Proteins; D011994:Recombinant Proteins; D020169:Caspases; C583895:type II metacaspase, tomato; D002118:Calcium
keywords: Activation; Arabidopsis metacaspase 1–9; AtMC1–9; Autoprocessing; Boc-Gly-Arg-Arg-MCA; GRR; LeMCA1; MCA2; Metacaspase; Norway spruce metacaspase; Site-directed mutagenesis; TaMCA4; Tomato; Trypanosoma brucei metacaspase 2; Yca1; Yeast metacaspase; mcII-Pa; tomato metacaspase; wheat metacaspase 4