Abstract
AT Pases were isolated from chloroplasts of the unicellular marine alga Acetabularia acetabulum. Two preparations of ATPase, a chloroplast-enriched fraction and an αβγ-complex were compared. The αβγ-complex was released into an EDTA solution and purified by anion-exchange chromatography, hydrophobic chromatography, and gel permeation chromatography. The subunit composition of this enzyme appeared to be 52-53 (α), 51(β), and 40(γ)kDa from SDS–PAGE. ATPase activity was enriched about 260-fold to a specific activity of approximate 4.1 U · mg protein −1. The catalytic properties of the αβγ-complex were as follows: pH optimum at 7.5; substrate specificity, ATP > ITP, GTP > UTP = CTP (Km for ATP 0.2 mM); divalent cation requirement, Mg2 + = Mn2 + = Co2 + > Zn2 + > Ni2 + > Ca2 +; ATPase activity was inhibited by monovalent anions, while monovalent cations had neither inhibitory nor stimulatory effects. Orthovanadate had no inhibitory effect on the enzyme activity of αβγ-complex. Azide was the most effective inhibitor of the αβγ-complex. N-Terminal amino acid sequences of the α and β subunits were not obtained and appeared to be blocked. The γ subunit gave a sequence of AGLKEMKD-XIGSVXNTKKI, which showed 60% similarity to the γ subunits of spinach and Chlamydomonas reinhardtii CF1-ATPase and EF1-ATPase.
| Original language | English |
|---|---|
| Pages (from-to) | 521-525 |
| Number of pages | 5 |
| Journal | Bioscience, Biotechnology and Biochemistry |
| Volume | 58 |
| Issue number | 3 |
| DOIs | |
| Publication status | Published - 1994 |
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ASJC Scopus subject areas
- Biotechnology
- Molecular Biology
- Biochemistry
- Organic Chemistry
- Analytical Chemistry
- Applied Microbiology and Biotechnology
- Food Science
- Biochemistry, Genetics and Molecular Biology(all)
- Chemistry (miscellaneous)
Cite this
Chloroplast atpase in acetabularia acetabulum : Purification and characterization of chloroplast f1-ATPase. / Satoh, Sachiko; Moritani, Chie; Oohashi, Toshitaka; Konishi, Kaori; Ikeda, Mikiko.
In: Bioscience, Biotechnology and Biochemistry, Vol. 58, No. 3, 1994, p. 521-525.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Chloroplast atpase in acetabularia acetabulum
T2 - Purification and characterization of chloroplast f1-ATPase
AU - Satoh, Sachiko
AU - Moritani, Chie
AU - Oohashi, Toshitaka
AU - Konishi, Kaori
AU - Ikeda, Mikiko
PY - 1994
Y1 - 1994
N2 - AT Pases were isolated from chloroplasts of the unicellular marine alga Acetabularia acetabulum. Two preparations of ATPase, a chloroplast-enriched fraction and an αβγ-complex were compared. The αβγ-complex was released into an EDTA solution and purified by anion-exchange chromatography, hydrophobic chromatography, and gel permeation chromatography. The subunit composition of this enzyme appeared to be 52-53 (α), 51(β), and 40(γ)kDa from SDS–PAGE. ATPase activity was enriched about 260-fold to a specific activity of approximate 4.1 U · mg protein −1. The catalytic properties of the αβγ-complex were as follows: pH optimum at 7.5; substrate specificity, ATP > ITP, GTP > UTP = CTP (Km for ATP 0.2 mM); divalent cation requirement, Mg2 + = Mn2 + = Co2 + > Zn2 + > Ni2 + > Ca2 +; ATPase activity was inhibited by monovalent anions, while monovalent cations had neither inhibitory nor stimulatory effects. Orthovanadate had no inhibitory effect on the enzyme activity of αβγ-complex. Azide was the most effective inhibitor of the αβγ-complex. N-Terminal amino acid sequences of the α and β subunits were not obtained and appeared to be blocked. The γ subunit gave a sequence of AGLKEMKD-XIGSVXNTKKI, which showed 60% similarity to the γ subunits of spinach and Chlamydomonas reinhardtii CF1-ATPase and EF1-ATPase.
AB - AT Pases were isolated from chloroplasts of the unicellular marine alga Acetabularia acetabulum. Two preparations of ATPase, a chloroplast-enriched fraction and an αβγ-complex were compared. The αβγ-complex was released into an EDTA solution and purified by anion-exchange chromatography, hydrophobic chromatography, and gel permeation chromatography. The subunit composition of this enzyme appeared to be 52-53 (α), 51(β), and 40(γ)kDa from SDS–PAGE. ATPase activity was enriched about 260-fold to a specific activity of approximate 4.1 U · mg protein −1. The catalytic properties of the αβγ-complex were as follows: pH optimum at 7.5; substrate specificity, ATP > ITP, GTP > UTP = CTP (Km for ATP 0.2 mM); divalent cation requirement, Mg2 + = Mn2 + = Co2 + > Zn2 + > Ni2 + > Ca2 +; ATPase activity was inhibited by monovalent anions, while monovalent cations had neither inhibitory nor stimulatory effects. Orthovanadate had no inhibitory effect on the enzyme activity of αβγ-complex. Azide was the most effective inhibitor of the αβγ-complex. N-Terminal amino acid sequences of the α and β subunits were not obtained and appeared to be blocked. The γ subunit gave a sequence of AGLKEMKD-XIGSVXNTKKI, which showed 60% similarity to the γ subunits of spinach and Chlamydomonas reinhardtii CF1-ATPase and EF1-ATPase.
UR - http://www.scopus.com/inward/record.url?scp=0028387615&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0028387615&partnerID=8YFLogxK
U2 - 10.1080/bbb.58.521
DO - 10.1080/bbb.58.521
M3 - Article
C2 - 7764691
AN - SCOPUS:0028387615
VL - 58
SP - 521
EP - 525
JO - Bioscience, Biotechnology and Biochemistry
JF - Bioscience, Biotechnology and Biochemistry
SN - 0916-8451
IS - 3
ER -