Give P's a chance
A Nobelist’s quest to open our eyes to the next DNA. It’s called Poly P
By SPYROS ANDREOPOULOS
For the past 15 years, Arthur Kornberg, MD, a Nobel Prize-winning biochemist at Stanford, has been deepening his relationship with one of biology’s wallflowers: a molecule he has nicknamed poly P. While most other biochemists ignore the omnipresent molecule (it shows up in every living cell on Earth) Kornberg, 89, can’t pull himself away. He’s convinced poly P is one of life’s great behind-the-scenes power players.
“It’s more important than I am,” he says, with a smile.
It’s not just his love of a good biochemical mystery that keeps the emeritus professor working in the lab — though that’s certainly part of what drives him. It’s also his conscience. With a little effort, he says, researchers could build on what he’s learned about this molecule to create revolutionary drugs for fighting infection.
Inorganic polyphosphate, the molecule Kornberg calls poly P, is older than the hills — literally. Phosphate rock and the hot, dry conditions that form poly P were present early on Earth before life forms developed. Scientists believe that the planet’s earliest cells likely exploited poly P’s unique molecular properties in their evolution. For example, poly P excels at shuttling useful metals such as iron and calcium into cells, and toxic metals such as cadmium and mercury out. And by sticking one of its easily detachable units onto an enzyme, poly P makes the enzyme water-soluble and as a result can completely change that enzyme’s function.
What exactly is poly P? It’s a chain of phosphate molecules. A single phosphate consists of a phosphorus atom surrounded by four oxygen atoms. In poly P, phosphates link to form chains — some up to thousands of units long.
Kornberg became fascinated by poly P in the 1950s. “We knew this polymer existed. It was found in a variety of cells, but we didn’t have the foggiest idea of what it did,” he remembers. A firm believer in the power of enzymes as research tools, Kornberg struck gold in 1956 when he and his colleagues found an enzyme that makes poly P from adenosine triphosphate — the main source of energy in cells. He dubbed the enzyme polyphosphate kinase, or PPK.
When Kornberg arrived at Stanford to lead the biochemistry department in the early ’60s, his work on DNA, for which he had received the Nobel Prize in 1959, had heated up. He shelved his interest in poly P to work on the enzymes responsible for DNA replication and repair, but he didn’t forget it. And in 1990, he was back at it.
One of his first insights was that while poly P is important for bacterial survival during good times — the organism’s growing phase — it’s even more important during harsh times — the stationary phase. When bacteria lacking PPK entered this phase, they died.
This underscored for him that the stationary phase of a microbial “bug” is anything but stationary. “More than 100 genes — probably several hundred genes — are turned on and off to give the cell the capacity to survive in the stationary phase,” Kornberg says. “The cell decides that things are going to get tough, and it makes adaptations. A bug in the soil or in sewage can survive as a species only by adapting to that condition so that when a nutrient comes along it can divide again. For lack of polyphosphate kinase, the bugs don’t express the genes needed to make adaptations, and they die.”
Kornberg’s lab went on to show that poly P increased the virulence of pathogenic bacteria responsible for many diseases. They include Bacillus anthracis, which causes anthrax; Salmonella, Shigella and Vibrio cholerae, sources of diarrheal dysenteries; the Neisseria that precipitate gonorrhea and meningitis; Pseudomonas aeruginosa, which produces the fatal pneumonia of cystic fibrosis and is the scourge of burn victims; Helicobacter pylori, the corrosive bug that incites peptic ulcers and stomach cancer; and Mycobacterium tuberculosis, responsible for afflicting millions with TB.
These inklings of poly P’s importance spurred Kornberg to conduct further investigations, which led to perhaps the most important finding of all, at least as far as drug development is concerned: Humans have their own version of PPK, but it’s different from the micro-organisms’ version.
And so it was that Kornberg’s curiosity about this chemical he’s seen referred to in textbooks as “a molecular fossil” led him to a potential solution to a huge real-world problem. Seen in total, his research suggests a new approach to combat disease-causing bacteria that have become resistant to available antibiotics. A drug targeting the bacterial PPK would knock out the bacteria but leave the human cells intact.
Further studies show that knocking out a pathogen’s poly P with PPK inhibitors delivers a one-two punch: increasing its vulnerability to attack by antibiotics and halting its toxin production.
This scenario is based in part on the role for poly P that’s finally emerging. In many bacteria, the molecule is a vital player in the organisms’ communication network. Without it, the network crashes. As a result, the bacteria produce lower levels of toxins, and they’re unable to form microbial communities called biofilms, which produce a thick, slimy layer that blocks antibiotics.
Kornberg knows that few others share his enthusiasm for poly P. Skeptics have looked at it as a mere laboratory curiosity, an attitude not uncommon when it comes to basic research. But the scientist thinks the amassing data will soon change critics’ minds. In 1970 his research on the role of DNA polymerase, a crucial enzyme relevant to DNA replication, also had its skeptics, he recalls, but they were proved wrong.
His lab’s initial studies using the bacterium P. aeruginosa offer encouragement. In one experiment, 14 of 15 mice infected with mutant bacteria that lacked poly P survived, whereas only one of 19 mice infected with normal bacteria carrying the PPK gene and poly P survived.
Promising. But because major drug companies are pulling out of antibiotic research, it’s no surprise that virtually no one is pursuing PPK inhibitors for drug development.
The exception is the progress made by Wenqing Xu, PhD, at the University of Washington together with Sam Lee, PhD, at the new biotech company Cocrystal Discovery Inc. Their crystallographic studies have refined the PPK binding sites in the bacteria P. aeruginosa and E. coli and have allowed them to model compounds that combine with the enzyme, potentially inhibiting its action. Like Kornberg, the scientists believe that PPK inhibitors have a chance to thwart antibiotic-resistant pathogens. Lee formed his Bothell, Wash., company this year to develop such drugs as well as antivirals.
Kornberg can count fewer than a dozen researchers who’ve pursued serious investigations of poly P: Igor Kulaev, PhD, at the Russian Academy of Sciences-Pushchino (dubbed by Kornberg as the prime “keeper of the torch”); Narayana Rao, PhD, Kornberg’s longtime collaborator at Stanford; Roberto Docampo, PhD, at the University of Georgia; Rosetta Reusch, PhD, at Michigan State University; and the crystallographic researchers Lee and Xu. Though their work adds to poly P’s profile, it’s not enough, Kornberg says.
So Kornberg continues his campaign for the molecule through scientific journals and through networking: in person, on the phone and online. “My fond hope is that knowledge of our work will influence other scientists and fields of investigation, spurred by our deep conviction that poly P is a molecule of many functions and not a fossil,” Kornberg says.
He’s undaunted by cessation of funding from the National Institutes of Health — like many established investigators, his support has disappeared during this time of NIH budget stagnation. And for the most part he’s unfazed by the widespread ignorance of the object of his fascination.
He continues speaking before groups of scientists and spreading the word among potential collaborators in academia and industry. His presentation to Stanford’s chemistry department was typical. Before the talk, none of the chemists had ever heard of the molecule. Disheartening, even for Kornberg. But the good news: Now they have.
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