One Rotary Mechanism for F^sub 1^-ATPase over ATP Concentrations from Millimolar down to Nanomolar
F^sub 1^-ATPase is a rotary molecular motor in which the central γ-subunit rotates inside a cylinder made of α^sub 3^β^sub 3^-subunits. The rotation is driven by ATP hydrolysis in three catalytic sites on the β-subunits. How many of the three catalytic sites are filled with a nucleotide during the c...
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Veröffentlicht in: | Biophysical journal 2005-03, Vol.88 (3), p.2047 |
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Sprache: | eng |
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Zusammenfassung: | F^sub 1^-ATPase is a rotary molecular motor in which the central γ-subunit rotates inside a cylinder made of α^sub 3^β^sub 3^-subunits. The rotation is driven by ATP hydrolysis in three catalytic sites on the β-subunits. How many of the three catalytic sites are filled with a nucleotide during the course of rotation is an important yet unsettled question. Here we inquire whether F^sub 1^ rotates at extremely low ATP concentrations where the site occupancy is expected to be low. We observed under an optical microscope rotation of individual F^sub 1^ molecules that carried a bead duplex on the γ-subunit. Time-averaged rotation rate was proportional to the ATP concentration down to 200 pM, giving an apparent rate constant for ATP binding of 2 × 10^sup 7^ M^sup -1^ S^sup -1^. A similar rate constant characterized bulk ATP hydrolysis in solution, which obeyed a simple Michaelis-Menten scheme between 6 mM and 60 nM ATP. F^sub 1^ produced the same torque of ~40 pN.nm at 2 mM, 60 nM, and 2 nM ATP. These results point to one rotary mechanism governing the entire range of nanomolar to millimolar ATP, although a switchover between two mechanisms cannot be dismissed. Below 1 nM ATP, we observed less regular rotations, indicative of the appearance of another reaction scheme. [PUBLICATION ABSTRACT] |
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ISSN: | 0006-3495 1542-0086 |