A novel cucumisin-like serine protease from leaf of legume Canavalia ensiformis

Autor: Raquel Elisa da Silva-López, Rayane Natashe Gonçalves, Dario E. Kalume, Maria Antonieta Ferrara
Rok vydání: 2020
Předmět:
Zdroj: Journal of Plant Biochemistry and Biotechnology. 30:147-159
ISSN: 0974-1275
0971-7811
Popis: Proteases are essential for plant physiology. Leguminosae species express high level of these enzymes, however, they were only reported in seeds. The present work isolated and characterized serine proteases of the aqueous extract from Canavalia ensiformis leaf (CE-A), a tropical legume. This extract was loaded on to benzamidine affinity column, and the serine protease fraction (CE-ABza) was purified 1.65-fold, yielding a total recovery of 62%. In a gelatin-SDS-PAGE, CE-ABza presented activity at 90 kDa under non-reducing, and 17, 32, and 90 kDa under reducing conditions. Peptidomimetic substrates for both trypsin and chymotrypsin as well as proteins with biotechnological relevance were digested, in distinctive levels, by CE-ABza. The maximal activity was at pH 8.5 and 9.5, and 40 °C. Protease activity was not affected at 70 °C for 24 h; however, it was completely inhibited by benzamidine and N-tosyl-L-phenylalanine chloromethylketone. Divalent cations had negative modulation on CE-ABza activity. Mass spectrometry experiments identified 11 orthologous proteases from this legume species, suggesting that CE-ABza shares similar and specific sequences especially with a serine protease, cucumisin. CE-ABza is a valuable source of very active and thermal stable serine proteases, which can be a potential candidate for biotechnological and therapeutical applications.
Databáze: OpenAIRE
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