Trypsin from jumbo squid (dosidicus gigas) hepatopancreas: Purification and characterization

Ana Gloria Villalba-Villalba, Enrique Márquez-Ríos, Marina Josafat Esquerra-Brauer, Francisco Javier Castillo-Yáñez

Research output: Contribution to journalArticlepeer-review

Abstract

Trypsin was purified from the hepatopancreas of Dosidicus gigas by fractionation with ammonium sulfate (30-70% saturation), gel filtration, affinity and ion exchange chromatography. The molecular weight of the trypsin obtained was ∼23.5kDa according to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and, showed a single band in the zymogram. The enzyme exhibited maximal activity at pH 8.5 and 40oC, using BAPNA as substrate. It was effectively inhibited by phenyl methyl sulfonyl fluoride (PMSF) (98%), and Na-p-Tosyl-L-lysine chloromethyl ketone (TLCK) (99%). Enzyme activity was affected by the following ions in decreasing order: Hg2+, Fe2+, Cu2+, Li1+, Mg2+, K1+, Mn2+, Ca2+. Trypsin activity decreased continuously as NaCl concentration increased from 0 to 30%. Km and kcat values were 0.085 ±1.45mM and 1.76 ±0.12s-1, respectively. The results suggest that the purified enzyme a potential agent to be used in biotechnological processes.
Original languageAmerican English
Pages (from-to)168-174
Number of pages7
JournalInterciencia
StatePublished - 1 Mar 2017
Externally publishedYes

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