I  n a vital chemical reaction within the membrane of the mitochondria—the powerhouse of the cell—a process called oxidative phosphorylation captures energy from the oxygen we breathe and the food we eat. During this process, the chemical adenosine triphosphate (ATP) is created; when the energy captured in ATP is used by cells, the ATP is split by the addition of water into adenosine diphosphate (ADP) and inorganic phosphate (Pi). The process had been recognized more than 50 years ago when Paul Boyer was a graduate student at the University of Wisconsin. But the key question remained: How did the reformation of ATP occur?

In the early 1950s, a researcher using an isotope of oxygen, 18O, discovered that mitochondria catalyzed a rapid exchange of Pi oxygens with water oxygens. Boyer, by now a faculty researcher at the University of Minnesota, began to use isotopes of oxygen and phosphorus (32P) in an effort to learn more about the oxidative phosphorylation process. He discovered that the overall process was dynamically reversible and that the 18O exchange was more rapid than the 32P exchange. This and other results led him and most others in the field to search for postulated intermediates in the formation of ATP. But efforts to find such intermediates were strikingly unsuccessful.

"We didn't understand the basis of our results at the time," Boyer recalls, smiling. "If we had, we would have been 20 years ahead." Instead, Boyer shifted his focus mostly to other enzymes, but continued some experiments on ATP formation.

By the 1970s, Boyer was at UCLA's interdepartmental Molecular Biology Institute, of which he was the founding director. One day while in a seminar, his mind wandered back to some unexplained oxygen-exchange data. A new concept for oxidative phosphorylation occurred to him. He realized that the intermediate he had searched for probably did not exist, and that an unexpected mechanism may occur.

"It became clear to me that the results could be explained if the energy from oxidations was not used to make the ATP molecule," he says, "but instead was used to bring about a release of a tightly bound ATP."

He had for the first time a satisfying insight into how the key enzyme, ATP synthase, makes the ATP that is used for a myriad of cellular processes, from muscle contractions to the transmission of nerve messages. Boyer estimates that an active person turns over a body weight of ATP in a day. "In a sense," he explains, "ATP serves as the currency of the cell." Plants, bacteria and other life forms use a similar ATP synthase; the formation of ATP appears to be the most prominent net chemical reaction occurring in the world. Boyer had synthesized a revolutionary concept for energy coupling.

But the new concept was not readily accepted. A paper submitted by Boyer to the leading publication in biochemistry, The Journal of Biological Chemistry, was declined.


 Skepticism abounded. But Boyer and his research associates felt that they were on the right track. Boyer exercised a perquisite of his recently received membership in the National Academy of Sciences—the ability to publish a paper without peer review—to forge ahead. "The concept just felt right," he explains. The paper was the beginning of what was to become known as the "binding change mechanism for ATP synthesis." The next decade of research by Boyer's group uncovered another unusual feature, the compulsory sequential participation of three catalytic subunits on the enzyme. By the early 1980s, many researchers in the field had accepted these aspects of the binding change mechanism.

But a third controversial concept, previously unknown in enzymology, was formulated by Boyer. Sophisticated 18O studies showed that the three catalytic sites behaved identically. The catalytic subunits appeared to be arranged in a circle around, and to be influenced by, an internal subunit. Boyer proposed that the internal piece behaves like a cam shaft in a car, rotating to cause the catalytic sites to go through three steps: ADP and Pi binding; tightening and ATP forming; and ATP releasing. During continuous rotational catalysis, all three sites would be in different conformations at any one time.

Thousands of enzymes had previously been studied, but none had shown even a hint of rotational catalysis. Boyer was out on a limb again. "This idea wasn't greeted with a lot of enthusiasm," he admits. Boyer's group obtained results supporting the concept of rotational catalysis, but lacked the technology for convincing assessment. By the late 1980s, Boyer, now 70, was easing into retirement. "I thought I would have to leave the problem of rotational catalysis undecided," he says.

E  nter John Walker, a senior scientist at the Medical Research Council Laboratory of Molecular Biology in Cambridge, England. In 1994, Walker and his colleagues reported painstaking X-ray crystallography data that defined the three-dimensional structure of the catalytic subunits of the ATP synthase. Walker's structure strikingly supported the concept of a rotational catalysis by establishing the predicted different conformations of the catalytic subunits and showing how the internal subunit could drive the conformational changes. Additional proof of rotational catalysis was shown by Masasuke Yoshida and colleagues in Japan, who provided a microscopically enhanced visual demonstration of the rotation.

The result: The 1997 Nobel Prize in Chemistry; Boyer and Walker shared a portion of the award for their studies on how ATP is formed, and Jens Skou of Denmark received a portion for his discovery of an enzyme in the brain and nerves that is one of the principal users of energy provided by ATP. This was a sweet moment of gratification for Boyer. It is the way of science, he notes, for skeptics to question and appraise new concepts. "I am fortunate that the enzyme I studied was not only very important, but had novel catalytic features," says Boyer. "It has been a privilege to be the first to recognize this remarkable molecular machine."

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