August 10, 2004 | David F. Coppedge

ATP Synthase: Another Unexpected Case of Fine Tuning

ATP synthase, the miniature rotary motor that powers our cells, has been a subject of great interest since the elucidation of its rotary function won three scientists a Nobel prize in 1997.  As an example of a precision-crafted, true electric rotary motor in living systems (another being the larger bacterial flagellum), it also provides a classic case study in intelligent design vs. evolution.  It has been the subject of frequent updates in these pages (start at 02/13/2004 and work backwards).  Now, another discovery about this ATP-synthesizing engine has revealed a deeper level of fine tuning.  Japanese scientists publishing in PNAS1 found a precision coupling between two components that was unexpected, yet apparently essential.
    For review, recall that ATP synthase has two functional domains, named F0 and F1.  The F1 part that actually synthesizes ATP from ADP + P is now fairly well understood.  It is composed of three pairs of lobes that spring-load ATP with every 120o turn of the camshaft, each pair of lobes either loading, catalyzing or ejecting an ATP molecule.  The F0 domain, however, has been harder to study.  Scientists knew it looks like a carousel of identical proteins, labeled c subunits.  Linked to it is a camshaft, named the gamma subunit, that drives the synthesis of ATP in F1.  Scientists knew the F0 carousel runs on protons delivered by a gumball-like mechanism named the a subunit (see 12/22/2003 headline).  But up till now, they were not sure how many c subunits comprised the carousel – or even if the number mattered.  Some studies had hinted that the F0 motor contained anywhere from 8 to 13 c subunits, depending on the species.  Now, the team of Mitome et al. found the answer: it is 10, and it must be 10 and only 10.  Other numbers don’t work.  That’s strange.  It means that F0 needs 10 protons per revolution, but F1 produces 3 ATP per revolution.  The ratio 10:3 is not an integer.  How can that be?
    The scientists arrived at the number 10 by customizing F0 rings with fixed numbers of c subunits, 2 through 14.  Then they linked them to the F1 domains and watched how much ATP was synthesized.  Results were obtained for only c=2, 5, and 10, which is interesting, considering that 2 and 5 are factors of 10.  The c=2 and c=5 cases produced a little ATP, and c=10 produced the maximum.  All the other numbers produced none.  The team deduced, therefore, that 10 (or one of its factors) is essential to match the proton-loading mechanism of the a subunit.
    The scientists also measured the proton flow through their custom carousels when disengaged from F1 and found, again, that 10 was the only number that worked.  Without 10 c subunits, no protons flowed.  Divide a circle of 360o by 10, and you get a 36o angle per c subunit during a complete revolution of the F0 motor.  The F1 domain, by contrast, produces ATP for each 120o turn, or 3 ATP per complete revolution.  The scientists seemed surprised that the proton-ATP ratio, “one of the most important parameters in bioenergetics,” is not an integer.  It’s as if three protons are sufficient to generate an ATP sometimes and four other times, because one cannot have a third of a proton.  Wouldn’t it be more logical if the number of c subunits was a multiple of three, say 6, 9, or 12?  With c=9, for instance, the camshaft angle would regularly line up with the F1 lobes every 3 protons, yielding one ATP every time, nice and neat.  The fact that it does not means that the coupling between F0 and F1 is not strict, as with toothed gears, but “permissive” – as if the two domains rotate according to their own structural needs, and are coupled together by a adaptor mechanism that has some degree of freedom to either twist or slip.
    The scientists ruled out slippage.  They knew that the camshaft can only produce an ATP in the F1 domain when it is lined up perfectly at the 120o steps.  Instead, they found that there is enough elastic flexibility in the camshaft to permit twist up to 40o during its rotation.  This flexibility allows the two domains to work separately, each according to its optimum configuration, with the twisting camshaft able to rock back and forth a little to give the F1 lobes time to complete their work.  In scientific lingo, “The flexibility of gamma allows both the F0-gamma and F1-gamma interfaces at the free-energy minima to stay in conformations adequate for the proton transport in F0 and the catalysis in F1 despite the step-size mismatch, providing sufficient time for those events to take place.”
    One more thing.  There isn’t much tolerance for error in this system.  The team found that a single point mutation at a spot named E56 in the c subunit was enough to quench all proton flow and all ATP synthesis: “This result provides evidence that each of all 10 E56 in the c-ring is indispensable.”  Also, the quantity of 10 subunits in the c-ring is critical, because 8, 9, 11, 12 and other numbers did not fit the gumball proton-delivery system of the a subunit: “Thus, the proton transport through F0 requires very strict arrangement of contact surface between F0-a and F0-c in the F0 assembly and even a rotary displacement as tiny as 3.3o (360o / 10 – 360o / 11) seems to be enough to disable a proton transfer between them.”
    The team made their measurements on ATP synthase motors from a species of thermophilic (heat-loving) bacteria.  They feel they have found a coupling strategy in living systems that could demonstrate a general principle: “Here, we report the permissive nature of the coupling between proton transport and ATP synthesis of F0-F1, but such nature of the coupling can be general among other biological motor systems to connect critical well tuned microscopic events in the large domain motions.”


1Mitome et al., “Thermophilic ATP synthase has a decamer c-ring: Indication of noninteger 10:3 H+/ATP ratio and permissive elastic coupling,” Proceedings of the National Academy of Sciences USA, 10.1073/pnas.0403545101, published online 8/09/04.

This discovery reveals a deeper level of design even more difficult to explain by evolution.  (As expected, these authors make no reference to evolution in their paper.)  A simple, easy-to-fathom machine would use the integer ratio; 3 protons yields one ATP.  The 10:3 ratio, puzzling at first, actually shows superior engineering.  It enables two disparate components with different operational requirements to be coupled together for the maximum efficiency of each.  In software, it would be like the driver that allows a device to work with any operating system.  In hardware, it would be like a tractor with a power-takeoff adapter that allows the engine to operate an attachment running at a different RPM.
      ATP synthase is made up of two finely tuned domains, F0 and F1, that operate under their own stringent requirements for function, but are useless without one another.  Why would the F0 c-ring carousel evolve by itself, if it had no function vital to the cell?  And how could the F1 system of six lobes, exquisitely-crafted for the synthesis of ATP, operate without an electrical motor to turn the camshaft?  The camshaft itself is a perfectly-designed component, with just the right amount of elastic flexibility, to couple the two very different domains.  Add to that the a subunit that feeds the protons at just the right rate and matches them to the appropriate active site on each c subunit, and the epsilon subunit that anchors the motor to the membrane, and you have an irreducibly complex system of irreducibly complex systems.  The fact that this whole composite machine works at near 100% efficiency is proof of product, a contrivance that virtually shouts “made by intelligent design.”

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