The aspartic proteinase is expressed in Arabidopsis thaliana seeds and localized in the protein bodies

Asuman Mutlu, Xia Chen, Sridhar M. Reddy, Susannah Gal

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

We have been studying a seed aspartic proteinase, termed AtAP, from Arabidopsis thaliana. In previous work, we purified the proteinase, analysed its activity and isolated the cDNA sequence. In this paper, the expression of the mRNA for the aspartic proteinase was analysed in seed tissues both by Northern blots for overall regulation and by in situ hybridization to follow cell-specific localization of message. We found a 1.9 kb aspartic proteinase message in dry seeds and seed pods. This message was expressed in many different cell types of the mature dry seed. The localization of the protein within these cells was also determined. Antibodies were raised against the AtAP and purified using affinity chromatography on an AtAP-immobilized-pepstatin A-agarose column. This purified antibody recognized several AtAP peptides in seeds. To localize the enzyme in cells, we isolated protein bodies from the dry seeds of Arabidopsis using a non-aqueous isolation method. The AtAP activity and antigenic peptides were found to be highest in the protein body fraction and colocalized with the seed storage protein 2S albumin and the vacuolar marker enzyme α-mannosidase. This protein body localization of the AtAP was confirmed with immunocytochemical localization by electron microscopy and shows that the protein is not secreted by these cells.

Original languageEnglish (US)
Pages (from-to)75-84
Number of pages10
JournalSeed Science Research
Volume9
Issue number1
StatePublished - Jan 1 1999

Fingerprint

aspartic proteinases
protein bodies
Arabidopsis thaliana
seeds
cells
mannosidases
peptides
antibodies
seed storage proteins
isolation techniques
affinity chromatography
enzymes
Northern blotting
in situ hybridization
agarose
albumins
pods
electron microscopy
proteinases
proteins

All Science Journal Classification (ASJC) codes

  • Plant Science

Cite this

@article{cb408f980f70458089a7bbdd82a212bb,
title = "The aspartic proteinase is expressed in Arabidopsis thaliana seeds and localized in the protein bodies",
abstract = "We have been studying a seed aspartic proteinase, termed AtAP, from Arabidopsis thaliana. In previous work, we purified the proteinase, analysed its activity and isolated the cDNA sequence. In this paper, the expression of the mRNA for the aspartic proteinase was analysed in seed tissues both by Northern blots for overall regulation and by in situ hybridization to follow cell-specific localization of message. We found a 1.9 kb aspartic proteinase message in dry seeds and seed pods. This message was expressed in many different cell types of the mature dry seed. The localization of the protein within these cells was also determined. Antibodies were raised against the AtAP and purified using affinity chromatography on an AtAP-immobilized-pepstatin A-agarose column. This purified antibody recognized several AtAP peptides in seeds. To localize the enzyme in cells, we isolated protein bodies from the dry seeds of Arabidopsis using a non-aqueous isolation method. The AtAP activity and antigenic peptides were found to be highest in the protein body fraction and colocalized with the seed storage protein 2S albumin and the vacuolar marker enzyme α-mannosidase. This protein body localization of the AtAP was confirmed with immunocytochemical localization by electron microscopy and shows that the protein is not secreted by these cells.",
author = "Asuman Mutlu and Xia Chen and Reddy, {Sridhar M.} and Susannah Gal",
year = "1999",
month = "1",
day = "1",
language = "English (US)",
volume = "9",
pages = "75--84",
journal = "Seed Science Research",
issn = "0960-2585",
publisher = "Cambridge University Press",
number = "1",

}

The aspartic proteinase is expressed in Arabidopsis thaliana seeds and localized in the protein bodies. / Mutlu, Asuman; Chen, Xia; Reddy, Sridhar M.; Gal, Susannah.

In: Seed Science Research, Vol. 9, No. 1, 01.01.1999, p. 75-84.

Research output: Contribution to journalArticle

TY - JOUR

T1 - The aspartic proteinase is expressed in Arabidopsis thaliana seeds and localized in the protein bodies

AU - Mutlu, Asuman

AU - Chen, Xia

AU - Reddy, Sridhar M.

AU - Gal, Susannah

PY - 1999/1/1

Y1 - 1999/1/1

N2 - We have been studying a seed aspartic proteinase, termed AtAP, from Arabidopsis thaliana. In previous work, we purified the proteinase, analysed its activity and isolated the cDNA sequence. In this paper, the expression of the mRNA for the aspartic proteinase was analysed in seed tissues both by Northern blots for overall regulation and by in situ hybridization to follow cell-specific localization of message. We found a 1.9 kb aspartic proteinase message in dry seeds and seed pods. This message was expressed in many different cell types of the mature dry seed. The localization of the protein within these cells was also determined. Antibodies were raised against the AtAP and purified using affinity chromatography on an AtAP-immobilized-pepstatin A-agarose column. This purified antibody recognized several AtAP peptides in seeds. To localize the enzyme in cells, we isolated protein bodies from the dry seeds of Arabidopsis using a non-aqueous isolation method. The AtAP activity and antigenic peptides were found to be highest in the protein body fraction and colocalized with the seed storage protein 2S albumin and the vacuolar marker enzyme α-mannosidase. This protein body localization of the AtAP was confirmed with immunocytochemical localization by electron microscopy and shows that the protein is not secreted by these cells.

AB - We have been studying a seed aspartic proteinase, termed AtAP, from Arabidopsis thaliana. In previous work, we purified the proteinase, analysed its activity and isolated the cDNA sequence. In this paper, the expression of the mRNA for the aspartic proteinase was analysed in seed tissues both by Northern blots for overall regulation and by in situ hybridization to follow cell-specific localization of message. We found a 1.9 kb aspartic proteinase message in dry seeds and seed pods. This message was expressed in many different cell types of the mature dry seed. The localization of the protein within these cells was also determined. Antibodies were raised against the AtAP and purified using affinity chromatography on an AtAP-immobilized-pepstatin A-agarose column. This purified antibody recognized several AtAP peptides in seeds. To localize the enzyme in cells, we isolated protein bodies from the dry seeds of Arabidopsis using a non-aqueous isolation method. The AtAP activity and antigenic peptides were found to be highest in the protein body fraction and colocalized with the seed storage protein 2S albumin and the vacuolar marker enzyme α-mannosidase. This protein body localization of the AtAP was confirmed with immunocytochemical localization by electron microscopy and shows that the protein is not secreted by these cells.

UR - http://www.scopus.com/inward/record.url?scp=0032933405&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0032933405&partnerID=8YFLogxK

M3 - Article

AN - SCOPUS:0032933405

VL - 9

SP - 75

EP - 84

JO - Seed Science Research

JF - Seed Science Research

SN - 0960-2585

IS - 1

ER -