June 15, 2005
Promiscuous Catalytic Activity Possessed by Novel Enzyme Structure
La Jolla, Calif.- Nature is a seemingly endless storehouse of interesting - and potentially life-saving - biological molecules. But tracking down and harvesting those chemicals in their natural form can be time-consuming, expensive and unreliable.
Now Salk scientists have discovered a new way of bringing "bio-prospecting" out of the rainforest and into the lab. Their findings are published in the June 16th edition of the journal Nature.
St©phane Richard, Joseph Noel and Tomohisa Kuzuyama isolated and examined a totally new enzyme that can mix and match biological chemicals to create a wide range of different molecules that could be used as the basis for new drugs. The enzyme, named Orf2, takes chemical building blocks known as small aromatic molecules and changes them by adding a fat-like molecule called a prenyl group. That modification can have a huge impact on where the aromatic molecule goes within the cell, and what sort of effects it has when it reaches its target.
"When you make that so-called hybrid molecule, all the chemistry associated with the original compound changes," said Noel. He explained that the addition of a prenyl group, also known as prenylation, produces a relatively large change in the chemical's biological properties. "The molecule becomes a little bit more greasy, and so the kinds of targets that it might interact with change completely."
"We actually have in our hands now the ability to do things that a chemist might not be able to do, or if a chemist was able to do it, it could be expensive and time-consuming," said Noel. "I think it's a completely novel way to approach the whole area of bio-prospecting and new natural medicines."
The group was interested in manipulating the properties of small aromatic molecules because these bacteria- and plant-derived compounds are involved in a wide variety of important biological processes. Some are potent cancer-fighting anti-oxidants, while others have antibiotic or anti-fungal properties. Many of these molecules only become active once they have had a prenyl group added.
The researchers tested whether their new enzyme could modify a wide range of small aromatic molecules, ranging from plant flavonoids - for instance, compounds found in hops that give beer its bitter taste or in other plants act as sunscreens, pigments, or antibiotics - to olivetol, a component in the production of THC, the active ingredient of marijuana. Unlike other previously known prenylation enzymes that only act on a small number of molecules, this new enzyme was able to attach prenyl groups to most of the different aromatic compounds tested. Some of the newly-formed hybrid molecules had previously been found in natural sources, but the enzyme also built some new compounds that had never been seen before.
The key to Orf2's flexibility seems to reside in the active site, the region of the enzyme where the small aromatic molecules bind. In many enzymes, the structure of the active site only allows one specific molecule to interact with it, like a key fitting into a lock. But the scientists found that Orf2's active site is a surprisingly spacious barrel-like structure, unlike any protein fold ever seen before. Not only does this wide barrel allow Orf2 to act on such a wide range of aromatic molecules, its discovery could also shed new light on the relationship between an enzyme's structure, the way it has evolved, and the chemical reactions it is able to carry out.
Although the researchers have not yet had a chance to test the modified molecules to see if their function has changed, they believe that Orf2 could be a powerful tool for creating biologically active compounds that could be used as drugs or as new methods of enhancing the disease-preventative properties of plants. Because the enzyme is able to modify such a wide range of compounds, scientists seeking to develop new drugs could use it in several different ways. If they had no particular end product in mind, they could feed interesting compounds to the enzyme and see what comes out. But researchers could also turn Orf2 into a more customized tool by using the techniques of genetic engineering to narrow its activity and tell it exactly which reactions to carry out.
"The big advantage that we have with our system is that it's a small, soluble, easy-to-work-with bacterial protein that seems to be able to do reactions that you can find somewhere else in nature, but where people have not been able yet to isolate the proteins," said St©phane Richard. "So it's kind of a short-cut - it's a way to bypass what is done already in nature, but for which we don't have the tools yet that nature has."
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