Monosaccharide composition and properties of a deglycosylated turnip peroxidase isozyme

Miguel A. Duarte-Vázquez, Blanca E. García-Almendárez, Arturo Rojo Dominguez, John R. Whitaker, C. Arroyave-Hernández, Carlos Regalado

Resultado de la investigación: Contribución a una revistaArtículo

17 Citas (Scopus)

Resumen

A neutral peroxidase isozyme (TP) purified from turnip (Brassica napus L. var. purple top white globe) was partially deglycosylated, using chemical and enzymatic treatment. A 32% carbohydrate removal was achieved by exposing TP to a mixture of PNGase F, O-glycosidase, NANase, GALase III and HEXase I, while m-periodate treatment removed about 88% of TP carbohydrate moiety. The glycoprotein fraction of the TP contained a relatively high mannose and fucose content (37 and 31%, w/w, respectively), 16% (w/w) galactose, and 15% (w/w) GlcNAc. Thus, the carbohydrate moiety was classified as a hybrid type. Partially deglycosylated TP had reduced activity (by 50-85%), was more susceptible to proteolysis, and showed a slight decrease in thermostability compared to the native enzyme. Circular dichroism studies strongly suggested that although the carbohydrate moiety of TP did not influence the conformation of the polypeptide backbone, its presence considerably enhanced protein conformational stability toward heat. Removal of oligosaccharide chains from TP caused a decrease in Km and Vmax for hydrogen peroxide. Native and chemically deglycosylated TP were similarly immunodetected by rabbit polyclonal antibodies raised against TP. The results suggest that the carbohydrate moiety of TP is important for peroxidase activity and stability.

Idioma originalInglés
Páginas (desde-hasta)5-11
Número de páginas7
PublicaciónPhytochemistry
Volumen62
N.º1
DOI
EstadoPublicada - 1 ene 2003

Huella dactilar

Brassica napus
turnips
Monosaccharides
monosaccharides
Peroxidase
Isoenzymes
isozymes
peroxidase
Carbohydrates
carbohydrates
Chemical analysis
Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Proteolysis
circular dichroism spectroscopy
heat stability
Fucose
fucose
Protein Stability
glycosidases
Glycoside Hydrolases

Citar esto

Duarte-Vázquez, M. A., García-Almendárez, B. E., Rojo Dominguez, A., Whitaker, J. R., Arroyave-Hernández, C., & Regalado, C. (2003). Monosaccharide composition and properties of a deglycosylated turnip peroxidase isozyme. Phytochemistry, 62(1), 5-11. https://doi.org/10.1016/S0031-9422(02)00456-9
Duarte-Vázquez, Miguel A. ; García-Almendárez, Blanca E. ; Rojo Dominguez, Arturo ; Whitaker, John R. ; Arroyave-Hernández, C. ; Regalado, Carlos. / Monosaccharide composition and properties of a deglycosylated turnip peroxidase isozyme. En: Phytochemistry. 2003 ; Vol. 62, N.º 1. pp. 5-11.
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title = "Monosaccharide composition and properties of a deglycosylated turnip peroxidase isozyme",
abstract = "A neutral peroxidase isozyme (TP) purified from turnip (Brassica napus L. var. purple top white globe) was partially deglycosylated, using chemical and enzymatic treatment. A 32{\%} carbohydrate removal was achieved by exposing TP to a mixture of PNGase F, O-glycosidase, NANase, GALase III and HEXase I, while m-periodate treatment removed about 88{\%} of TP carbohydrate moiety. The glycoprotein fraction of the TP contained a relatively high mannose and fucose content (37 and 31{\%}, w/w, respectively), 16{\%} (w/w) galactose, and 15{\%} (w/w) GlcNAc. Thus, the carbohydrate moiety was classified as a hybrid type. Partially deglycosylated TP had reduced activity (by 50-85{\%}), was more susceptible to proteolysis, and showed a slight decrease in thermostability compared to the native enzyme. Circular dichroism studies strongly suggested that although the carbohydrate moiety of TP did not influence the conformation of the polypeptide backbone, its presence considerably enhanced protein conformational stability toward heat. Removal of oligosaccharide chains from TP caused a decrease in Km and Vmax for hydrogen peroxide. Native and chemically deglycosylated TP were similarly immunodetected by rabbit polyclonal antibodies raised against TP. The results suggest that the carbohydrate moiety of TP is important for peroxidase activity and stability.",
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Duarte-Vázquez, MA, García-Almendárez, BE, Rojo Dominguez, A, Whitaker, JR, Arroyave-Hernández, C & Regalado, C 2003, 'Monosaccharide composition and properties of a deglycosylated turnip peroxidase isozyme', Phytochemistry, vol. 62, n.º 1, pp. 5-11. https://doi.org/10.1016/S0031-9422(02)00456-9

Monosaccharide composition and properties of a deglycosylated turnip peroxidase isozyme. / Duarte-Vázquez, Miguel A.; García-Almendárez, Blanca E.; Rojo Dominguez, Arturo; Whitaker, John R.; Arroyave-Hernández, C.; Regalado, Carlos.

En: Phytochemistry, Vol. 62, N.º 1, 01.01.2003, p. 5-11.

Resultado de la investigación: Contribución a una revistaArtículo

TY - JOUR

T1 - Monosaccharide composition and properties of a deglycosylated turnip peroxidase isozyme

AU - Duarte-Vázquez, Miguel A.

AU - García-Almendárez, Blanca E.

AU - Rojo Dominguez, Arturo

AU - Whitaker, John R.

AU - Arroyave-Hernández, C.

AU - Regalado, Carlos

PY - 2003/1/1

Y1 - 2003/1/1

N2 - A neutral peroxidase isozyme (TP) purified from turnip (Brassica napus L. var. purple top white globe) was partially deglycosylated, using chemical and enzymatic treatment. A 32% carbohydrate removal was achieved by exposing TP to a mixture of PNGase F, O-glycosidase, NANase, GALase III and HEXase I, while m-periodate treatment removed about 88% of TP carbohydrate moiety. The glycoprotein fraction of the TP contained a relatively high mannose and fucose content (37 and 31%, w/w, respectively), 16% (w/w) galactose, and 15% (w/w) GlcNAc. Thus, the carbohydrate moiety was classified as a hybrid type. Partially deglycosylated TP had reduced activity (by 50-85%), was more susceptible to proteolysis, and showed a slight decrease in thermostability compared to the native enzyme. Circular dichroism studies strongly suggested that although the carbohydrate moiety of TP did not influence the conformation of the polypeptide backbone, its presence considerably enhanced protein conformational stability toward heat. Removal of oligosaccharide chains from TP caused a decrease in Km and Vmax for hydrogen peroxide. Native and chemically deglycosylated TP were similarly immunodetected by rabbit polyclonal antibodies raised against TP. The results suggest that the carbohydrate moiety of TP is important for peroxidase activity and stability.

AB - A neutral peroxidase isozyme (TP) purified from turnip (Brassica napus L. var. purple top white globe) was partially deglycosylated, using chemical and enzymatic treatment. A 32% carbohydrate removal was achieved by exposing TP to a mixture of PNGase F, O-glycosidase, NANase, GALase III and HEXase I, while m-periodate treatment removed about 88% of TP carbohydrate moiety. The glycoprotein fraction of the TP contained a relatively high mannose and fucose content (37 and 31%, w/w, respectively), 16% (w/w) galactose, and 15% (w/w) GlcNAc. Thus, the carbohydrate moiety was classified as a hybrid type. Partially deglycosylated TP had reduced activity (by 50-85%), was more susceptible to proteolysis, and showed a slight decrease in thermostability compared to the native enzyme. Circular dichroism studies strongly suggested that although the carbohydrate moiety of TP did not influence the conformation of the polypeptide backbone, its presence considerably enhanced protein conformational stability toward heat. Removal of oligosaccharide chains from TP caused a decrease in Km and Vmax for hydrogen peroxide. Native and chemically deglycosylated TP were similarly immunodetected by rabbit polyclonal antibodies raised against TP. The results suggest that the carbohydrate moiety of TP is important for peroxidase activity and stability.

KW - Brassica napus L. var. purple top white globe

KW - Chemical deglycosylation

KW - Cruciferae

KW - Enzymatic deglycosylation

KW - Turnip peroxidase

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U2 - 10.1016/S0031-9422(02)00456-9

DO - 10.1016/S0031-9422(02)00456-9

M3 - Artículo

C2 - 12475613

AN - SCOPUS:0038601418

VL - 62

SP - 5

EP - 11

JO - Phytochemistry

JF - Phytochemistry

SN - 0031-9422

IS - 1

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Duarte-Vázquez MA, García-Almendárez BE, Rojo Dominguez A, Whitaker JR, Arroyave-Hernández C, Regalado C. Monosaccharide composition and properties of a deglycosylated turnip peroxidase isozyme. Phytochemistry. 2003 ene 1;62(1):5-11. https://doi.org/10.1016/S0031-9422(02)00456-9