Proc Natl Acad Sci U S A 2002 Feb 19;99(4):1882-7
Forced engagement of a RNA/protein complex by a chemical inducer of dimerization to modulate gene expression.
Harvey I, Garneau P, Pelletier J.
Department of Biochemistry and McGill Cancer Center, McIntyre Medical Sciences Building, McGill University, Montreal, QC, Canada H3G 1Y6.
A general strategy is described for forcing the engagement of an RNA/protein complex by using small-molecule ligands. A bivalent molecule was created by linking a protein-binding ligand to an RNA-binding ligand. On presentation of the chemical inducer of dimerization to the RNA by the protein, cooperative binding ensued, resulting in higher-affinity complexes. When the chemical inducer of dimerization was used to target the protein to an mRNA template, the resulting RNA/protein complex was sufficiently stable to inhibit mRNA translation. This approach provides a logic to modulate gene expression by using small-molecule ligands to recruit protein surfaces to mRNAs.
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from today's BioCentury (thanks, R!).........
“The flexible nature of RNA allows it to change shape,
making it difficult to design small molecules that will bind and
remain bound with high affinity. Also, many binding pockets
either appear or disappear depending on what proteins are
bound to the RNA at a given time,” said Vincent Stanton Jr., vice
president and principal scientist at Variagenics Inc.
Researchers at McGill University in Montreal avoided this
problem by creating a chemical inducer of dimerization (CID), a
bivalent molecule that recognizes mRNA at a specific sequence
that also binds to a specific protein, in essence gluing the mRNA
and proteins together.
Specifically, the researchers showed that a tobramycin-based
CID caused a 5- to 10-fold reduction in the production of
reporter protein in vitro. Though their work is the first peer
reviewed publication using the CID technology, VGNX (Cambridge,
Mass.) filed a European patent application covering the
CID methodology in 2000. The McGill researchers subsequently
initiated CID research at the company’s suggestion, though
VGNX did not fund the study.
The binding constant of the RNA/CID/protein complex was
in the nanomolar range, which is much better than the affinity of
tobramycin for the RNA itself, showing that the CID’s contribution
to the complex is critical. Without it, no protein/RNA
complex would exist at all.
The simultaneous binding of the CID to the protein and
mRNA puts the two in close proximity, allowing the RNA to not
only bind to the CID, but to wrap around the protein as well.
“The cooperativity in binding is extremely important because
it attenuates the ability of the mRNA to bind to the
ribosome,” Stanton told BioCentury. The article showed that the
tobramycin-based CID inhibited translation by disrupting the
formation of the 80S ribosome, which is necessary for proper
protein translation.
The strategy theoretically could allow small molecules to be
designed to disrupt the production of any protein, but Stanton
noted that the application of CIDs towards therapeutics is still in
development. “It is clear that this will not work with every
mRNA/protein pair and it will be necessary to identify cellular
proteins that interact cooperatively with RNA and develop CIDs
that bind with a high affinity to them,” he said.
Stanton added that though VGNX is doing CID research, the
company’s major focus remains pharmacogenomics.
(doesn't sound like it's high priority stuff) |
