3/1/01

Dawn Levy, News Service (650) 725-1944; e-mail: dawnlevy@stanford.edu

Genetic engineering speeds development of new antibiotics

Our world is full of all kinds of bacteria--the good, the bad and the innocuous. Most often, it's the bad bacteria that catch our attention with their health-stealing antics. But sometimes, as the old adage goes, it takes a thief to catch a thief.

By hijacking the biosynthetic machinery of bacteria, scientists can create antibiotics to kill the bad bacteria that rob us of our vitality. Now, genetic engineers in the Stanford lab of Professor Chaitan Khosla have inserted the largest working genes to date into the Escherichia coli bacterium, transforming this run-of-the-mill microbe into an organism that can churn out new precursors of erythromycin, a broad-spectrum antibiotic and penicillin substitute. Their feat, to be published in the Mar. 2 issue of Science magazine, demonstrates a powerful tool for developing novel antibiotics to combat bacteria that have become resistant to overused antibiotics.

Traditionally, manufacturers make erythromycin commercially through fermentation using the soil bacterium Saccharopolyspora erythraea. The process is hard to scale up, creating a bottleneck in the drug-development process.

To modify erythromycin and give it novel properties, chemists start with pure erythromycin and use chemistry to change the molecule--a costly approach. "If life is at stake, that expense is worthwhile," says Khosla, a professor of chemistry and chemical engineering. "In this experiment, instead of using chemistry, we reprogram genes to make a modified erythromycin. It's a more efficient way to do genetic engineering than had ever been done before."

Ten years ago, Khosla and colleagues asked: Can one harness nature's biosynthetic machinery to make erythromycin? The group's success in using genetically engineered bacteria to make the antibiotic ­ a process Khosla calls "doing chemistry by genetics" ­ proves the answer is yes.

"Historically, bacteria have been great sources of new pharmaceuticals," Khosla says. "Drugs have been isolated from all sorts of weird sources in nature and modified." In nature, bacteria may produce antibiotics to inhibit the growth of nearby strains that compete for nutritional resources.

The genetic engineers have "tweaked nature's strategies" to make antibiotics with novel properties, Khosla says: "The machinery is highly maleable and can be manipulated to make modified natural products." Modifications may make antibiotics better able to combat resistant strains of bacteria. Other variations may confer therapeutic properties besides antibiotic functions.

Stanford graduate student Blaine Pfeifer and postdoctoral researcher Suzanne Admiraal did the project's experimental work. Hugo Gramajo of the Facultad de Ciencias Bioquimicas y Farmaceuticas in Argentina provided a substance required to make a key cellular substrate. David Cane of Brown University served as an additional adviser on the project, which was supported with grants from the National Institutes of Health and the National Science Foundation.

Pfeifer started with E. coli, deleting four native genes and introducing six foreign genes. Several genes ­ each over 10,000 base pairs in size ­ had never before been functionally expressed in E. coli.

"We didn't know a priori if we'd even be able to have genes that big expressed in E. coli," Pfeifer says. "We didn't know if E. coli would behave like the native host (S. erythraea) which produces the natural antibiotic."

But the genetically engineered E. coli worked as well as the bacterium that makes erythromycin in nature: It began cranking out the amino-acid building blocks encoded by the different genes. The amino acids were placed one after another, like pearls in a necklace, to form proteins. The proteins folded into active shapes, and chemical groups were added to the proteins to confer unique properties. And proteins were made in the right amounts and at the proper speeds for efficient assembly of the antibiotic precursor. In nature, the same process happens in S. erythraea.

But S. erythraea grows slowly ­ a population of this bacterial strain takes four hours to double in number. A population of E. coli, in contrast, only takes 20 minutes to double. That, plus the fact that a lot is already known about E. coli, used extensively in bioscience research, makes the latter the workhorse of choice in genetic engineering.

For now, genetically engineered E. coli "give us a whole new toolbox" for building new antibiotics, Pfeifer says. But in the near future, Khosla adds, this Clark-Kent-turned-Superman microbe may serve as a practical means of manufacturing new life-saving drugs.

-30-

By Dawn Levy