Chloroplast ATPase in A cetabularia acetabulum

ATPases 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...

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Published in:Bioscience, biotechnology, and biochemistry biotechnology, and biochemistry, 1994-03, Vol.58 (3), p.521
Main Authors: Satoh, Sachiko, Moritani, Chie, Ohhashi, Toshitaka, Konishi, Kaori, Ikeda, Mikiko
Format: Article
Language:English
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Summary:ATPases 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+ = Mn = Co2+ > Zn2+ > Ni2+ > Ca2+ ; ATPase activity was inhibited by monovalent anions (NO-3, SCN-), 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 βsubunits 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.
ISSN:0916-8451
1347-6947