The catalytic site of the pectin biosynthetic enzyme α-1,4-galacturonosyltransferase is located in the lumen of the Golgi

J. D. Sterling, H. F. Quigley, A. Orellana, D. Mohnen

Resultado de la investigación: Article

97 Citas (Scopus)

Resumen

α-1,4-Galacturonosyltransferase (Ga1AT) is an enzyme required for the biosynthesis of the plant cell wall pectic polysaccharide homogalacturonan (HGA). Ga1AT activity in homogenates from pea (Pisum sativum L. var. Alaska) stem internodes co-localized in linear and discontinuous sucrose gradients with latent UDPase activity, an enzyme marker specific for Golgi membranes. Ga1AT activity was separated from antimycin A-insensitive NADH:cytochrome c reductase and cytochrome c oxidase activities, enzyme markers for the endoplasmic reticulum and the mitochondria, respectively. Ga1AT and latent UDPase activities were separated from the majority (80%) of callose synthase activity, a marker for the plasma membrane, suggesting that little or no Ga1AT is present in the plasma membrane. Ga1AT activities in proteinase K-treated and untreated Golgi vesicles were similar, whereas no Ga1AT activity was detected after treating Golgi vesicles with proteinase K in the presence of Triton X-100. These results demonstrate that the catalytic site of Ga1AT resides within the lumen of the Golgi. The products generated by Golgi-localized Ga1AT were converted by endopolygalacturonase treatment to mono- and di-galacturonic acid, thereby showing that Ga1AT synthesizes 1→4-linked α-D-galacturonan. Our data provide the first enzymatic evidence that a glycosyltransferase involved in HGA synthesis is present in the Golgi apparatus. Together with prior results of in vivo labeling and immunocytochemical studies, these results show that pectin biosynthesis occurs in the Golgi. A model for the biosynthesis of the pectic polysaccharide HGA is proposed.

Idioma originalEnglish
Páginas (desde-hasta)360-371
Número de páginas12
PublicaciónPlant Physiology
Volumen127
N.º1
DOI
EstadoPublished - 1 ene 2001

Huella dactilar

active sites
pectins
Catalytic Domain
biosynthesis
Endopeptidase K
Peas
Enzymes
plasma membrane
polysaccharides
enzymes
1,3-beta-glucan synthase
enzyme activity
Polysaccharides
antimycin A
glycosyltransferases
galacturonic acid
NADH dehydrogenase
Cell Membrane
polygalacturonase
Antimycin A

ASJC Scopus subject areas

  • Physiology
  • Genetics
  • Plant Science

Citar esto

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title = "The catalytic site of the pectin biosynthetic enzyme α-1,4-galacturonosyltransferase is located in the lumen of the Golgi",
abstract = "α-1,4-Galacturonosyltransferase (Ga1AT) is an enzyme required for the biosynthesis of the plant cell wall pectic polysaccharide homogalacturonan (HGA). Ga1AT activity in homogenates from pea (Pisum sativum L. var. Alaska) stem internodes co-localized in linear and discontinuous sucrose gradients with latent UDPase activity, an enzyme marker specific for Golgi membranes. Ga1AT activity was separated from antimycin A-insensitive NADH:cytochrome c reductase and cytochrome c oxidase activities, enzyme markers for the endoplasmic reticulum and the mitochondria, respectively. Ga1AT and latent UDPase activities were separated from the majority (80{\%}) of callose synthase activity, a marker for the plasma membrane, suggesting that little or no Ga1AT is present in the plasma membrane. Ga1AT activities in proteinase K-treated and untreated Golgi vesicles were similar, whereas no Ga1AT activity was detected after treating Golgi vesicles with proteinase K in the presence of Triton X-100. These results demonstrate that the catalytic site of Ga1AT resides within the lumen of the Golgi. The products generated by Golgi-localized Ga1AT were converted by endopolygalacturonase treatment to mono- and di-galacturonic acid, thereby showing that Ga1AT synthesizes 1→4-linked α-D-galacturonan. Our data provide the first enzymatic evidence that a glycosyltransferase involved in HGA synthesis is present in the Golgi apparatus. Together with prior results of in vivo labeling and immunocytochemical studies, these results show that pectin biosynthesis occurs in the Golgi. A model for the biosynthesis of the pectic polysaccharide HGA is proposed.",
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The catalytic site of the pectin biosynthetic enzyme α-1,4-galacturonosyltransferase is located in the lumen of the Golgi. / Sterling, J. D.; Quigley, H. F.; Orellana, A.; Mohnen, D.

En: Plant Physiology, Vol. 127, N.º 1, 01.01.2001, p. 360-371.

Resultado de la investigación: Article

TY - JOUR

T1 - The catalytic site of the pectin biosynthetic enzyme α-1,4-galacturonosyltransferase is located in the lumen of the Golgi

AU - Sterling, J. D.

AU - Quigley, H. F.

AU - Orellana, A.

AU - Mohnen, D.

PY - 2001/1/1

Y1 - 2001/1/1

N2 - α-1,4-Galacturonosyltransferase (Ga1AT) is an enzyme required for the biosynthesis of the plant cell wall pectic polysaccharide homogalacturonan (HGA). Ga1AT activity in homogenates from pea (Pisum sativum L. var. Alaska) stem internodes co-localized in linear and discontinuous sucrose gradients with latent UDPase activity, an enzyme marker specific for Golgi membranes. Ga1AT activity was separated from antimycin A-insensitive NADH:cytochrome c reductase and cytochrome c oxidase activities, enzyme markers for the endoplasmic reticulum and the mitochondria, respectively. Ga1AT and latent UDPase activities were separated from the majority (80%) of callose synthase activity, a marker for the plasma membrane, suggesting that little or no Ga1AT is present in the plasma membrane. Ga1AT activities in proteinase K-treated and untreated Golgi vesicles were similar, whereas no Ga1AT activity was detected after treating Golgi vesicles with proteinase K in the presence of Triton X-100. These results demonstrate that the catalytic site of Ga1AT resides within the lumen of the Golgi. The products generated by Golgi-localized Ga1AT were converted by endopolygalacturonase treatment to mono- and di-galacturonic acid, thereby showing that Ga1AT synthesizes 1→4-linked α-D-galacturonan. Our data provide the first enzymatic evidence that a glycosyltransferase involved in HGA synthesis is present in the Golgi apparatus. Together with prior results of in vivo labeling and immunocytochemical studies, these results show that pectin biosynthesis occurs in the Golgi. A model for the biosynthesis of the pectic polysaccharide HGA is proposed.

AB - α-1,4-Galacturonosyltransferase (Ga1AT) is an enzyme required for the biosynthesis of the plant cell wall pectic polysaccharide homogalacturonan (HGA). Ga1AT activity in homogenates from pea (Pisum sativum L. var. Alaska) stem internodes co-localized in linear and discontinuous sucrose gradients with latent UDPase activity, an enzyme marker specific for Golgi membranes. Ga1AT activity was separated from antimycin A-insensitive NADH:cytochrome c reductase and cytochrome c oxidase activities, enzyme markers for the endoplasmic reticulum and the mitochondria, respectively. Ga1AT and latent UDPase activities were separated from the majority (80%) of callose synthase activity, a marker for the plasma membrane, suggesting that little or no Ga1AT is present in the plasma membrane. Ga1AT activities in proteinase K-treated and untreated Golgi vesicles were similar, whereas no Ga1AT activity was detected after treating Golgi vesicles with proteinase K in the presence of Triton X-100. These results demonstrate that the catalytic site of Ga1AT resides within the lumen of the Golgi. The products generated by Golgi-localized Ga1AT were converted by endopolygalacturonase treatment to mono- and di-galacturonic acid, thereby showing that Ga1AT synthesizes 1→4-linked α-D-galacturonan. Our data provide the first enzymatic evidence that a glycosyltransferase involved in HGA synthesis is present in the Golgi apparatus. Together with prior results of in vivo labeling and immunocytochemical studies, these results show that pectin biosynthesis occurs in the Golgi. A model for the biosynthesis of the pectic polysaccharide HGA is proposed.

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