Cells get two chances, not just one, to fix their mistakes

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Cells get two chances, not just one, to fix their mistakes

http://researchnews.osu.edu/archive/cellerror.htm

COLUMBUS, Ohio – Cells have two chances to fix the same mistake in their

protein-making process instead of just one – a so-called proofreading step –

that had previously been identified, according to new research.

Proteins are essential to life and do most of the work in cells, so avoiding

mistakes during their production is a critical way to prevent a variety of

harmful conditions that result when mutations cause damage or when cells die.

Better knowledge of the mechanism behind these occasional mistakes could

increase understanding of various disease processes, especially in

neurodegenerative disorders, some of which scientists suspect are associated

with mutated proteins, Ohio State University researchers say.

The discovery of this second step also gives drug-makers a new target to

consider, especially in the development of antibiotics. Drugs designed to

interfere with the enzymes that make, find and repair the mistakes during

protein production could be powerful agents in stopping bacterial cell growth.

One key enzyme involved in cell quality control is called phenylalanyl-tRNA

synthetase (PheRS). This enzyme's job within the cell is to correctly select one

of the amino acids that will be strung together into a molecule to make a

protein.

" We're describing a pretty simple process where the cell says, `I think I'll

have one more look at that,' " said Ibba, senior author of the study and

an associate professor of microbiology at Ohio State. " It looks at the building

blocks and checks that they're right before it makes the protein. "

The research is published in the March 13 issue of the journal Molecular Cell.

In past examinations of this mechanism in the cell, Ibba's lab had identified a

single quality-control measure cells take as they produce proteins. The

researchers initially thought that this proofreading step was the only check

during the protein-making process.

" There was a step at which we thought, now it's done, and if a mistake gets

through here, it's irreversible and is going to end up as a mistake. But it

turns out there is yet another step at which the cell has another look. It's

checking itself, " Ibba said. " The bottom line is we must have been missing

something. "

Previous research has suggested that cells, on average, make one error during

protein production for every 10,000 amino acids strung together.

" What we do is try to find out where that error rate number comes from, " said

Ibba, also an investigator in Ohio State's Biochemistry Program and its Center

for RNA Biology. " Understanding where the mistakes come from means you can try

to predict conditions that will either raise or lower the frequency of

mistakes. "

Within the cell, PheRS is one member of a family of enzymes responsible for

selecting amino acids that will be attached to an adapter molecule that

facilitates the protein-building process. The amino acids must be attached to

the appropriate adapter to ensure the genetic code is deciphered properly.

Ibba's lab has been studying this enzyme's activities for years. After observing

over time what appeared to be a second quality-control step, the scientists had

to devise a method they could use to prove the second step occurred. So the

researchers first generated the mistake synthetically, and then introduced other

enzymes that would normally be present later in protein production to see if

they could then observe the second quality check.

The researchers discovered that the same enzyme that makes the mistake, PheRS,

also checks and cleans up after itself in a process that removes the incorrect

amino acid and attaches the correct one in its place. And the enzyme can do this

even after an initial check misses the mistake and allows the protein-building

process to continue.

" The enzyme is two catalysts, one that can make the mistake and one that can

correct the mistake. It can let the mistake go and grab it back. Nothing tells

it to do this. It figures it out on its own, " Ibba said.

These experiments were conducted using E. coli bacterial cells, which are a

preferred model for many cell studies. But understanding this mechanism can be

particularly useful in the design of antibiotics because many such drugs

specifically target the protein-production process to halt the growth of

bacteria.

" We're trying to understand the process which in the past has proven to be very

fruitful as a target for antibiotics, " Ibba said. " The hope is when you target

protein synthesis in general, either you stop it completely or make the process

too inaccurate so the cell can't grow. "

This very same quality-control process, involving a different enzyme, is being

targeted in the development of an antifungal agent that is currently being

tested in humans to treat toenail fungus, Ibba noted.

Even with this second editing step identified, there is still plenty to learn.

For example, these enzymes do not act alone. Their interactions with other

enzymes in the cell affect their behavior. And exactly what happens when mutant

proteins slip through the quality-control system remains poorly understood, as

well.

" Sometimes mistakes do get in, and that's what we're still uncertain about. Even

in some neurodegenerative disorders, we can see that there are almost certainly

errors, but the frequency is impossible to know at present, " Ibba said. " If we

know more about the mechanism, then if we find mutations, we'll have a much

better chance of finding what the consequences of those mutations are. "

This research was funded primarily by the National Science Foundation, with

additional support from the National Institutes of Health and the American Heart

Association.

All of the study's investigators are affiliated with Ohio State's Center for RNA

Biology. Co-authors on the study are Jiqiang Ling of the Biochemistry Program;

Byung Ran So in chemistry; Srujana Yadavalli, Herve Roy and Shinichiro Shoji in

microbiology; Kurt Fredrick of the Biochemistry Program and microbiology; and

Karin Musier-Forsyth of the Biochemistry Program and the departments of

chemistry and biochemistry.