RT Journal Article
T1 Glyoxyl-ficin: an example where a more intense multipoint covalent attachment may decrease enzyme stability
A1 Siar, El-Hocine
A1 Morellon-Sterling, Roberto
A1 Carballares, Diego
A1 Rocha Martín, Javier
A1 Barbosa, Oveimar
A1 Bolívar Bolívar, Juan Manuel
A1 Fernandez-Lafuente, Roberto
AB The immobilization of ficin extract (sulfidryl proteases) on glyoxyl agarose beads stabilized it, but there is a clear stability maximum after 3 h of immobilization, being the 24 h biocatalyst less stable when thermally inactivated in presence of air. The incubation of the thermally inactivated biocatalysts in the presence of dithiothreitol permitted to recover almost total activity of the biocatalysts prepared for 24 h of immobilization, while the enzyme immobilized for 3 h recovered less activity. This suggested that the main cause of the faster enzyme inactivation of the 24 h biocatalyst was the Cys oxidation. This was confirmed by inactivating the Cys blocked biocatalysts by using 2,2′-dipyridyldisulfide. Again, a higher percentage of the initial activity (when unblocked by incubation in mercaptoethanol the thermally inactivated biocatalysts) was observed for the 24 h biocatalyst. The use of helium atmosphere enabled to greatly improve the enzyme stability of all immobilized biocatalysts, confirming the highest stability of the biocatalyst immobilized for 24 h. Results suggested that this biocatalyst was in fact the most rigid one, but that the distortion induced by the immobilization produced a higher exposition of the catalytic Cys, and this way, its oxidation becomes faster in the presence of oxygen.
PB Elsevier
SN 1359-5113
YR 2023
FD 2023
LK https://hdl.handle.net/20.500.14352/101622
UL https://hdl.handle.net/20.500.14352/101622
LA eng
NO Siar E-H, Morellon-Sterling R, Carballares D, Rocha-Martin J, Barbosa O, Bolivar JM, et al. Glyoxyl-ficin: an example where a more intense multipoint covalent attachment may decrease enzyme stability. Process Biochemistry. 2023;132:289-96
NO We gratefully recognize the financial support from Ministerio de Ciencia e Innovacion and Agencia Estatal de Investigacion  (Spanish Government) (PID2021-122398OB-I00 and PID2022-136535OB-I00). The authors also thank the financial support from Ministerio de Ciencia, Tecnología e Innovacion, Ministerio de Educacion Nacional, Ministerio de Industria, Comercio y Turismo, and ICETEX, Programme Ecosistema Científico-Colombia Científica from Fondo Francisco Jose de Caldas; Grant RC-FP44842-212-2018. RMS thanks the Ministerio de Educacion -Spanish Government for a FPU fellowship, DC thank to Ministerio de Ciencia e Innovacion-Spanish ´ Government for a FPI. Prof A. Berenguer (Alicante University) help and suggestions during the writing of this paper are gratefully recognized.
NO Ministerio de Ciencia e Innovación (España)
NO Ministerio de Educacion (España)
NO Ministerio de Educación (Colombia)
DS Docta Complutense
RD 10 abr 2025