Functional Incorporation of Chimeric b Subunits into F sub(1)F sub(o) ATP Synthase

• Published in: Journal of bacteriology Vol. 189; no. 15; pp. 5463 - 5471
• Main Authors: Claggett, Shane B, Grabar, Tammy Bohannon, Dunn, Stanley D, Cain, Brian D
• Format: Journal Article
• Language: English
• Published: 01-08-2007
• Subjects: Escherichia coli; Thermosynechococcus elongatus
• ISSN: 0021-9193; 1098-5530

F sub(1)F sub(o) ATP synthases function by a rotary mechanism. The enzyme's peripheral stalk serves as the stator that holds the F sub(1) sector and its catalytic sites against the movement of the rotor. In Escherichia coli, the peripheral stalk is a homodimer of identical b subunits, but photosynthetic bacteria have open reading frames for two different b-like subunits thought to form heterodimeric b/b' peripheral stalks. Chimeric b subunit genes have been constructed by substituting sequence from the Thermosynechococcus elongatus b and b' genes in the E. coli uncF gene, encoding the b subunit. The recombinant genes were expressed alone and in combination in the E. coli deletion strain KM2 ( Delta b). Although not all of the chimeric subunits were incorporated into F sub(1)F sub(o) ATP synthase complexes, plasmids expressing either chimeric b sub(E39-I86) or b' sub(E39-I86) were capable of functionally complementing strain KM2 ( Delta b). Strains expressing these subunits grew better than cells with smaller chimeric segments, such as those expressing the b' sub(E39-D53) or b sub(L54-I86) subunit, indicating intragenic suppression. In general, the chimeric subunits modeled on the T. elongatus b subunit proved to be more stable than the b' subunit in vitro. Coexpression of the b sub(E39-I86) and b' sub(E39-I86) subunits in strain KM2 ( Delta b) yielded F sub(1)F sub(o) complexes containing heterodimeric peripheral stalks composed of both subunits.