Antibodies to the F₁‐ATPase of Rhodospirillum rubrum and Its Purified Native β‐Subunit:

Abstract

1 . Antibodies prepared against the Rhodospirillunt ruhrum F1-ATPase (RrFI) and its purified, native [hubunit, exhibited cross-reactivity with thc following soluble preparations of R. ruhrunt ATPase: RrFo . FI, RrFl and thc 8-subunit. Anti-RrF1, but not anti-l) antibodies, also formed precipitin lines with soluble @-less RrFl, indicating that antigenic determinants of both the 1-subunit and the other four RrFl-subunits are expressed in the whole RrFl molecule. Both antibodies agglutinated the R. rubvum chromatophores, suggesting that the [hubunit is located on the external part of RrF1.2. Both antibodies inhibited ATP synthesis and hydrolysis activities of R. rubrum chromatophores, as well a s all the soluble ATPase reactions. Similar concentrations of each antibody were required for 50 inhibition of all these reactions, but anti-RrFI was always somewhat more effective than anti-P. These data indicate that the []-subunit is involved in the catalytic site of the RrFl-enzyme.3. The antibodies prepared against R. rubrum F1-ATPase and its P-subunit could bind thc soluble chloroplast F1-ATPase (CF1) and inhibited ATP-linked reactions carried out by chloroplasts and by soluble CFI. In these reactions, unlike in the R. rubrun? ones, anti-@ was a more potent inhibitor than the anti-RrF1 antibody.The cross-reaction obtained between the antibodies raised against R. rubrurn F1 and its b-subunit and the chloroplast CF1 indicates the presence of similar antigenic determinants in the photosynthetic prokaryotic and eukaryotic F1-ATPases, which have been conserved during evolution.

Hybrid Rhodospirillum rubrumF0F1 ATP Synthases Containing Spinach Chloroplast F1 β or α and β Subunits Reveal the Essential Role of the α Subunit in ATP Synthesis and Tentoxin Sensitivity

“…One such system, where both ATP synthesis and hydrolysis can be followed, was obtained by LiCl treatment of chromatophores isolated from the photosynthetic bacterium Rhodospirillum rubrum (12). This treatment was found to release the bulk of their ␤ subunits (12,13) resulting in the loss of over 90% of their ATP synthesis and hydrolysis activities. Both activities could be restored upon reconstituting the treated chromatophores with the released subunits.…”

The photosynthetic F1-?3?3 and ?1?1 catalytic core complexes

“…Both p h o t o s y n t h e t i c CFl-O~3fl3 and RrFl-C~l/31 complexes have however been released in an assembled form from the membrane-bound CFoFI and RrFoFl by extraction with 2 M LiC1 in presence of 4 mM MgATP (Avital and Gromet-Elhanan 1991; Andralojc and Harris 1992). This technique was developed for the selective release of all the RrFlfl subunit from the chromatophore membrane-bound RrFoF1 (Philosophet al 1977;Philosoph and Gromet-Elhanan 1981), but could release also between 5 to 20% of the RrFl a (Andralojc and Harris 1993;Gromet-Elhanan 1995). It involves a 30 min extraction at 4 °C followed by 6 h centrifugation at 200000 g, which is required for removal of all the extracted chromatophores (Gromet-Elhanan and Khananshvili 1986).…”

Purification and Properties of the Coupling‐Factor ATPases F1 from Rhodopseudomonas palustris and Rhodopseudomonas sphaeroides

“…SDS polyacrylamide gradient gel electrophoresis of dissociated RpFl and RsFl revealed five components for each enzyme, denoted a, p, y, 6, and E in order of increasing electrophoretic mobilities. The electrophoretic pattern obtained from dissociated RpFl is illustrated in Fig.…”

“…Coupling-factor ATPases are composed of two structurally and functionally distinct sectors, the hydrophilic F1 displaying ATP hydrolase activity and the hydrophobic Fo mediating H ' translocation across the membrane. In agreement with the abbreviations CF1 and RrFl for the F1-ATPases from chloroplasts and R. rubrum [6], we denoted the corresponding proteins from Rhodopseudomonas palustris RpFl and from R. sphaeroides RsFl.Although it is now realized that the energy-transducing membranes of chloroplasts and bacteria contain similar ATPase systems, there are still some differences regarding solubilization of the ATPase from the membranes [3,7-101, latency and unmasking of the ATPase activity [1,2,5,12], cation specificity [I, 5,10,12], substrate specificity [I, 131, cold or heat lability [I, 51, molecular weight [4,5,14] plus subunit composition [15,16], and kinetic properties [5,12,13,17].Recently, we have reported on the solubilization of an ATPase protein from R. palustris [18]. The crude ATPase…”

“…Coupling-factor ATPases are composed of two structurally and functionally distinct sectors, the hydrophilic F1 displaying ATP hydrolase activity and the hydrophobic Fo mediating H ' translocation across the membrane. In agreement with the abbreviations CF1 and RrFl for the F1-ATPases from chloroplasts and R. rubrum [6], we denoted the corresponding proteins from Rhodopseudomonas palustris RpFl and from R. sphaeroides RsFl.…”