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In very rare cases - typically a fraction of a
percent - transfer RNA binds with the wrong
amino acid. If this error, or mistranslation, is not corrected, that
mistranslated amino acid is ultimately incorporated into a protein.
Past research by the Schimmel team and others has shown that as little
as one wrong amino acid can have profound consequences for health.
"Even such a tiny defect can overwhelm a cell's ability to deal with
misfolded proteins, ultimately causing specific neurological problems,"
says Scripps Research molecular biologist Kirk Beebe, first author of
the new paper with Marissa Mock.
Quality Control
The prevailing thinking among researchers in the field has been that
the portions of synthetases that recognize and bind transfer RNA and
appropriate amino acids are so accurate that little editing is
required. What little editing does occur was thought tied to the same
checkpoints within the enzymes that perform recognition and binding.
But the current study suggests a new perspective is needed, at least
for one widely studied enzyme, a synthetase found in everything from
bacteria to humans that binds the amino acid alanine.
The Schimmel team's research revealed that a completely distinct
segment of the enzyme acts as a second checkpoint responsible for
identifying mistranslations and removing any amino acid besides
alanine that might attach to the alanine transfer RNA. Remarkably,
this second zone within the enzyme focuses its activity on the very
same two nucleotides in the genetic code of the transfer RNA used by
the first checkpoint, a guanine and uracil pair referred to as G3�U70.
"The part that is astonishing is that the information that each of
these checkpoints is looking for is embedded in the same transfer RNA
molecule," says Schimmel, "There's no precedent for this that we're
aware of."
The researchers were able to show that this editor checkpoint, even
when separated from the rest of the enzyme, was able to efficiently
cleave mistranslated amino acids from the alanine transfer RNA.
Further experiments revealed that when the G3�U70 pair was transferred
to a different type of transfer RNA, the editing unit still removed a
non-alanine amino acid, showing clearly that the pair is the trigger
for the activity.
Mystery Solved?
The research also led to a surprising find regarding segments of
genome-encoded fragments common in cells and sometimes referred to as
freestanding domains, whose functions have been debated for many years.
The team found that certain freestanding domains with a genetic
sequence very similar to the second checkpoint in the alanine
synthetase can independently remove mistranslated amino acids in test
tube experiments. This suggests that the fragments may act as yet
another checkpoint to ensure that proteins are properly synthesized.
The activity by the freestanding domain, called AlaXp, also targets
the G3�U70 pair.
Whether AlaXp actually edits within cells is not certain, and how they
could perform such a function is also not yet clear. "That's still a
question to be resolved," says Beebe, "But our work presents a great
possibility for how things are likely to occur." The group has already
begun new experiments to study AlaXp activity in mouse cells.
"The results of the study were a complete surprise to us," says
Schimmel.
Beebe and Schimmel agree that the apparent triple redundancy the team
discovered for preventing mistranslation highlights the importance of
accurate protein synthesis. It is unlikely such a robustly redundant
system would have evolved, they say, if this were not the case.
The work's impact should go beyond adding to the limited understanding
of how cells avoid errors during protein synthesis. "We do think there
are probably powerful connections to disease," says Schimmel.
Schimmel's and other groups have already shown clear connections
between individual and cumulative errors in translation and a variety
of problems tied to aging, such as neurodegenerative conditions. Now
the Schimmel team has begun collaborative work with a number of other
research groups to identify in synthetases telltale genetic mutations
referred to as small nucleotide polymorphisms that can lead to
mistranslation. Identifying such errors could reveal the underlying
causes of diseases currently not understood and possibly identify
potential paths for treatment.
In addition to Beebe, Mock, and Schimmel, Eve Merriman was an author
on the new Nature paper, entitled "Distinct domains of tRNA synthetase
recognize the same base pair." All authors are from The Scripps
Research Institute.
The study was funded by the National Institutes of Health, the Skaggs
Foundation, and the National Foundation for Cancer Research.
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