Formation and properties of hybrid photosynthetic F1-ATPases

Demonstration of different structural requirements for stimulation and inhibition by tentoxin

Ward C. Tucker, Ziyun Du, Zippora Gromet-Elhanan, Mark L. Richter

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

A hybrid ATPase composed of cloned chloroplast ATP synthase β and γ subunits (βC and γC) and the cloned α subunit from the Rhodospirillum rubrum ATP synthase (αR) was assembled using solubilized inclusion bodies and a simple single-step folding procedure. The catalytic properties of the assembled α3 Rβ3 Cγ C were compared to those of the core α3 Cβ3 Cγ C complex of the native chloroplast coupling factor 1 (CF1) and to another recently described hybrid enzyme containing R. rubrum α and β subunits and the CF1 γ subunit (α3 Rβ3 Rγ C). All three enzymes were similarly stimulated by dithiothreitol and inhibited by copper chloride in response to reduction and oxidation, respectively, of the disulfide bond in the chloroplast γ subunit. In addition, all three enzymes exhibited the same concentration dependence for inhibition by the CF1 ε subunit. Thus the CF1 γ subunit conferred full redox regulation and normal ε binding to the two hybrid enzymes. Only the native CF1 α3 Cβ3 Cγ C complex was inhibited by tentoxin, confirming the requirement for both CF1 α and β subunits for tentoxin inhibition. However, the α3 R β3 CγC complex, like the α3 Cβ3 Cγ C complex, was stimulated by tentoxin at concentrations in excess of 10 μM. In addition, replacement of the aspartate at position 83 in βC with leucine resulted in the loss of stimulation in the α3 Rβ3 Cγ C hybrid. The results indicate that both inhibition and stimulation by tentoxin require a similar structural contribution from the β subunit, but differ in their requirements for α subunit structure.

Original languageEnglish (US)
Pages (from-to)2179-2186
Number of pages8
JournalEuropean Journal of Biochemistry
Volume268
Issue number7
DOIs
StatePublished - Sep 27 2001

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Proton-Translocating ATPases
Demonstrations
Chloroplast Proton-Translocating ATPases
Rhodospirillum rubrum
Enzymes
Dithiothreitol
Inclusion Bodies
Chloroplasts
Aspartic Acid
Leucine
Disulfides
Oxidation-Reduction
Adenosine Triphosphatases
Chlorides
Copper
Adenosine Triphosphate
Oxidation
tentoxin

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Formation and properties of hybrid photosynthetic F1-ATPases : Demonstration of different structural requirements for stimulation and inhibition by tentoxin. / Tucker, Ward C.; Du, Ziyun; Gromet-Elhanan, Zippora; Richter, Mark L.

In: European Journal of Biochemistry, Vol. 268, No. 7, 27.09.2001, p. 2179-2186.

Research output: Contribution to journalArticle

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