נגישות
menu      
חיפוש מתקדם
תחביר
חפש...
הספר "אוצר וולקני"
אודות
תנאי שימוש
ניהול
קהילה:
אסיף מאגר המחקר החקלאי
פותח על ידי קלירמאש פתרונות בע"מ -
ATP-dependent association between subunits of Clp protease in pea chloroplasts
Year:
2001
Source of publication :
Planta
Authors :
אוסטרזצר, אורן
;
.
Volume :
213
Co-Authors:
Halperin, T., Department of Agricultural Botany, The Hebrew University of Jerusalem, Rehovot 76100, Israel
Ostersetzer, O., Department of Agricultural Botany, The Hebrew University of Jerusalem, Rehovot 76100, Israel
Adam, Z., Department of Agricultural Botany, The Hebrew University of Jerusalem, Rehovot 76100, Israel
Facilitators :
From page:
614
To page:
619
(
Total pages:
6
)
Abstract:
The chloroplast ATP-dependent Clp protease (EC 3.4.21.92) is composed of the proteolytic subunit ClpP and the regulatory ATPase, ClpC. Although both subunits are found in the stroma, the interaction between the two is dynamic. When immunoprecipitation with antibodies against ClpC was performed on stroma from dark-adapted pea (Pisum sativum L. cv. Alaska) chloroplasts, ClpC but not ClpP was precipitated. However, when stroma was supplemented with ATP, both ClpC and ClpP were precipitated. Co-immunoprecipitation was even more efficient in the presence of ATP-γ-S, suggesting that the association between regulatory and proteolytic subunits is dependent on binding of ATP to ClpC, but not its hydrolysis. To further test this association, stroma was fractionated by column chromatography, and the presence of Clp subunits in the different fractions was monitored immunologically. When stroma depleted of ATP was fractionated on an ion-exchange column, ClpP and ClpC migrated separately, whereas in the presence of ATP-γ-S both subunits co-migrated. Similar results were observed in size-exclusion chromatography. To further characterize the precipitated enzyme, its proteolytic activity was assayed by testing its ability to degrade β-casein. No degradation was observed in the absence of ATP, and degradation was inhibited in the presence of phenylmethylsulfonyl fluoride, consistent with Clp being an ATP-dependent serine protease. The activity of the isolated enzyme was further tested using chimeric OE33 as a model substrate. This protein was also degraded in an ATP-dependent manner, supporting the suggested role of Clp protease as a major housekeeping protease in the stroma.
Note:
Related Files :
beta casein
chlorophyll
enzymes
immunoprecipitation
Pisum sativum
עוד תגיות
תוכן קשור
More details
DOI :
10.1007/s004250100527
Article number:
Affiliations:
Database:
סקופוס
Publication Type:
מאמר
;
.
Language:
אנגלית
Editors' remarks:
ID:
24595
Last updated date:
02/03/2022 17:27
Creation date:
17/04/2018 00:08
Scientific Publication
ATP-dependent association between subunits of Clp protease in pea chloroplasts
213
Halperin, T., Department of Agricultural Botany, The Hebrew University of Jerusalem, Rehovot 76100, Israel
Ostersetzer, O., Department of Agricultural Botany, The Hebrew University of Jerusalem, Rehovot 76100, Israel
Adam, Z., Department of Agricultural Botany, The Hebrew University of Jerusalem, Rehovot 76100, Israel
ATP-dependent association between subunits of Clp protease in pea chloroplasts
The chloroplast ATP-dependent Clp protease (EC 3.4.21.92) is composed of the proteolytic subunit ClpP and the regulatory ATPase, ClpC. Although both subunits are found in the stroma, the interaction between the two is dynamic. When immunoprecipitation with antibodies against ClpC was performed on stroma from dark-adapted pea (Pisum sativum L. cv. Alaska) chloroplasts, ClpC but not ClpP was precipitated. However, when stroma was supplemented with ATP, both ClpC and ClpP were precipitated. Co-immunoprecipitation was even more efficient in the presence of ATP-γ-S, suggesting that the association between regulatory and proteolytic subunits is dependent on binding of ATP to ClpC, but not its hydrolysis. To further test this association, stroma was fractionated by column chromatography, and the presence of Clp subunits in the different fractions was monitored immunologically. When stroma depleted of ATP was fractionated on an ion-exchange column, ClpP and ClpC migrated separately, whereas in the presence of ATP-γ-S both subunits co-migrated. Similar results were observed in size-exclusion chromatography. To further characterize the precipitated enzyme, its proteolytic activity was assayed by testing its ability to degrade β-casein. No degradation was observed in the absence of ATP, and degradation was inhibited in the presence of phenylmethylsulfonyl fluoride, consistent with Clp being an ATP-dependent serine protease. The activity of the isolated enzyme was further tested using chimeric OE33 as a model substrate. This protein was also degraded in an ATP-dependent manner, supporting the suggested role of Clp protease as a major housekeeping protease in the stroma.
Scientific Publication
You may also be interested in