Every time he peers into Nature's voluminous bag of biochemical tricks, Craig Townsend, professor of chemistry, comes away amazed.
"Nature is absolutely the master of organic chemists," Townsend says. "There's a lot to learn from the master."
Townsend, postdoctoral fellow Rong-Feng Li and graduate student Tony Stapon have been taking an "apprenticeship" from streptomyces and erwinia, families of bacteria. The topic of study is production of a prized class of antibiotics, carbapenems, originally created by these bacteria.
In results published in the Journal of the American Chemical Society last month, Townsend's group announced that they'd found that erwinia make carbapenems by using three unusual and impressively efficient reactions, which do the chemical equivalent of pushing a large rock up a hill. That same metaphorical rock later takes a downhill plunge that gives the carbapenems their germ-"smashing" power.
"One practical application of this might someday be the ability to produce some of these antibiotics more effectively," Townsend says. "A better understanding of the natural way bacteria produce these compounds could, for example, lead to less expensive solutions for their synthesis."
Carbapenems belong to a larger class of antibiotics known as beta-lactams. The distinguishing characteristic of a beta-lactam is a high-energy ring of three carbon atoms and one nitrogen atom known as a beta-lactam ring.
Many well-known beta-lactams, including penicillin and cephalosporin, have become vulnerable to antibiotic resistance. Beginning in the 1950s and 1960s, an increasing number of bacteria started producing beta-lactamases, enzymes that rob beta-lactams of their germ-killing power.
Carbapenems, though, have remained unaffected by most beta-lactamases, making them an important tool for controlling resistant infections.
"The problem is that carbapenems are naturally produced by bacteria only in very small amounts," Townsend explains. Most other antibiotics are made by fermentation, but carbapenems aren't currently amenable to those processes. That leaves synthesis as the only option for commercial carbapenem production, greatly increasing the cost of the drugs.
Townsend's group has worked to understand how various species of bacteria make the four known classes of carbapenems. They also studied bacterial production of carbapenams, a closely related family of compounds with no antibiotic properties.
"It's been a long process using a variety of techniques," he explains. "We've had to use everything from molecular biology all the way to organic chemistry and synthesis of potential precursors to isolate and characterize the products. It's just the kind of thing I like to see happening in bio-organic chemistry, that a question that isn't easily addressable from any single discipline can be attacked and solved using techniques from many different areas of science."
Experiments in Cambridge, England, recently linked carbapenem production to a cluster of genes in one of the bacteria. By disrupting individual genes in the cluster and by transplanting them to bacteria that do not normally produce carbapenems, researchers found that five of nine genes in the cluster were required for the production of carbapenems.
"We asked ourselves, Using these genes to make predictions on how some of the reactions' proteins will appear, can we make any analogies to known enzyme reactions? Are the functions or active sites in the proteins involved similar or different, and can we use that information to identify other compounds involved in the reactions?" Townsend says.
However, Townsend's group generally found that the proteins are either unfamiliar or the bacteria used them in unexpected ways.
"The role of the enzymes, if you were to look at just the gene sequences and the translated proteins, was not so obvious," Townsend recalls. "There are enzymes and enzyme reactions that are understood from studies of primary metabolism and basic biochemistry that have been changed. They have evolved in these bacteria to doing some other kind of task."
The chemically toughest of those other tasks is locking up potential energy in the intermediate and final stages of carbapenam production. Chemical reactions tend to favor release of energy and typically create reaction products with less potential energy than the materials that went into the reaction. The intermediate and final stages of carbapenem production, though, are "thermodynamically uphill"--they lock more potential energy into the reaction products.
"Things have to happen in each of these reactions to make the desired results for carbapenem production energetically favorable," Townsend says. "So the enzymes use co-factors, they use molecular oxygen, they use other things so that the overall reaction becomes thermodynamically possible. And perhaps the biggest surprise for us was that for each of the four known classes of beta-lactams, the bacteria's solutions for achieving these goals were different."
Energy bound up in the carbapenems leaves the beta-lactam ring wanting to pop open, but it typically doesn't do that until another microbial enzyme comes along and tries to use it in a reaction. When that happens, the ring reacts with the protein, fatally disabling enzyme and bacteria alike.
In a parallel effort, Townsend's lab is studying the production of clavulanic acid, a material produced by bacteria that combats bacterial resistance to beta-lactams.
"Finding ways to improve old antibiotics and finding strategies to create new antibiotics is an important task for medicinal chemistry and organic chemistry, because microbial resistance is a problem that's always going to be with us," he says.