News and Publications
The Skaggs Institute For Chemical Biology
Scientific Report 1997-1998
Decoding Genetic Information in Translation
P. Schimmel, R. Alexander, J. Chihade, C. Fabrega, M. Farrow, T. Hendrickson,
A. Morales, B. Nordin, E. O'Donoghue, T. Nomanbhoy, L. Ribas de Pouplana, N.
Sardesai, B. Steer, M. Swairjo, R. Turner, K. Wakasugi, C.-C. Wang
The genetic code was established more than 2 billion years ago and became
universally part of all living organisms. The rules of the code, which relate
specific nucleotide triplets to specific amino acids, are determined by aminoacylation
reactions catalyzed by aminoacyl tRNA synthetases. In these reactions, an amino
acid is associated with a specific nucleotide triplet of the genetic code by
virtue of being linked to a specific tRNA that harbors the anticodon triplet
cognate to the amino acid. Because of their central role in establishing the
rules of the code, the tRNAs are thought to have arisen quite early, perhaps
in the context of an RNA world. The synthetases may have been among the earliest
proteins to appear, perhaps replacing ribozymes that catalyzed the aminoacylation
of primordial tRNAs.
We wish to understand all aspects of these systems. Areas of research include
detailed studies of molecular recognition of RNA, as represented by the synthetase-tRNA
interactions and peptide models of those interactions; editing and correction
of potential errors in translation, as represented by the misacylation of tRNAs
and misactivation of amino acids; reconstruction of the assembly of tRNA through
domain-domain interactions of RNA pieces; and dissection of catalytic and RNA-binding
components of tRNA synthetases. We are also interested in evolutionary relationships
between synthetases that can reveal insight into the origins of the code and
protein synthesis; involvement of eukaryotic synthetases in higher order cellular
functions, such as cell signaling pathways and the construction of macromolecular
and design and synthesis of systems of tRNA-like molecules that can be acylated
and assemble complexes for peptide synthesis, as examples of the potential for
early systems of protein synthesis that predated the ribosome.
The cloverleaf structure of tRNA is folded into 2 domains (Fig. 1). One domain
contains the anticodon with the template-reading head of the genetic code; the
other, called the minihelix domain, contains the amino acid attachment site.
The minihelix domain itself is a substrate for aminoacylation for at least 10
of the synthetases. Pieces even smaller than the minihelix are active as substrates
(Fig. 2). Because these pieces lack the anticodon trinucleotides, the relationship
between the sequences and structures of these active pieces (with as few as 4
bp) and the specific amino acid is distinct from the genetic code. The relationship
between sequences and structures in tRNA acceptor stems and specific amino acids
is referred to as an operational RNA code for amino acids. The operational RNA
code is thought to have predated the genetic code.
A scheme for the assembly of the synthetase-tRNA complex in evolution is
given in Figure 3. Each domain of the synthetase interacts with a separate domain
of the tRNA. The minihelix domain of the tRNA contains the amino acid attachment
site and is thought to be the earliest version of a tRNA molecule. The anticodon-containing
domain was added later in evolution. Similarly, the catalytic domain of the synthetase
is thought to be the historical enzyme, which contains the active site and which
interacts with the minihelix domain of the tRNA. Later, a second domain of the
synthetase was added; this domain interacts with the second domain of the tRNA.
This model is useful for guiding experiments aimed at reconstructing the assembly
of the synthetase-tRNA complexes in evolution.
Our recent studies have expanded on these concepts to elucidate a role for
the tRNA in fine-structure recognition of amino acids. That is, the ability to
distinguish between 2 closely similar amino acids is greatly enhanced by an effector
function of the tRNA, which causes the rejection of amino acids that are not
exactly matched with the synthetase and its cognate tRNA. This RNA-dependent
fine-structure recognition may have developed first in an RNA world and later
was incorporated into the synthetase system as a critical part of maintaining
the accuracy of the genetic code.
Alexander, R., Nordin, B., Schimmel, P. Activation of microhelix charging
by localized helix destabilization. Proc. Natl. Acad. Sci. U.S.A. 95:12214, 1998.
Beuning, P.J., Yang, F., Schimmel, P., Musier-Forsyth, K. Specific
atomic groups and RNA helix geometry in acceptor stem recognition by a tRNA synthetase.
Proc. Natl. Acad. Sci. U.S.A. 94:10150, 1997.
Chihade, J., Hayashibara, K., Shiba, K., Schimmel, P. Strong selective
pressure to mark an RNA acceptor stem for alanine. Biochemistry 37:9193, 1998.
Frugier, M., Schimmel, P. Subtle atomic group recognition in the RNA
minor groove. Proc. Natl. Acad. Sci. U.S.A. 94:11291, 1997.
Glasfeld, E., Schimmel, P. Zinc-dependent tRNA binding by a peptide
element within a tRNA synthetase. Biochemistry 37:6739, 1997.
Hale, S.P., Auld, D.S., Schmidt, E., Schimmel, P. Discrete nucleotides
in tRNA for editing and aminoacylation. Science 276:1250, 1997.
Hale, S.P., Schimmel, P. DNA aptamer targets translational editing
motif in a tRNA synthetase. Tetrahedron 53:11985, 1997.
Henderson, B.S., Beuning, P.J., Shi, J.-P., Bald, R., Fürste, J.P.,
Erdmann, V.A., Musier-Forsyth, K., Schimmel, P. Subtle functional interactions
in the RNA minor groove at a nonessential base pair. J. Am. Chem. Soc. 120:9110,
Henderson, B.S., Schimmel, P. RNA-RNA interactions between oligonucleotide
substrates for aminoacylation. Bioorg. Med. Chem. 5:1071, 1997.
Musier-Forsyth, K., Schimmel, P. Atomic determinants for aminoacylation
of RNA minihelices and relationship to genetic code. Accounts Chem. Res., in
Nair, S., Ribas de Pouplana, L., Houman, F., Avruch, A., Shen, X., Schimmel,
P. Species-specific tRNA recognition in relation to tRNA synthetase contact
residues. J. Mol. Biol. 269:1, 1997.
Nureki, O., Vassylyev, D.G., Tateno, M., Shimada, A., Nakama, T., Fukai,
S., Konno, M., Hendrickson, T.L., Schimmel, P., Yokoyama, S. Enzyme structure
with two catalytic sites for double-sieve selection of substrate. Science 280:578,
Ribas de Pouplana, L., Beuchter, D., Sardesai, N.Y., Schimmel, P. Functional
analysis of peptide motif for RNA microhelix binding suggests new family of RNA-binding
domains. EMBO J. 17:5449, 1998.
Ribas de Pouplana, L., Schimmel, P. Reconstruction of quaternary structures
of class II tRNA synthetases by rational mutagenesis of a conserved domain. Biochemistry
Ribas de Pouplana, L., Turner, R.J., Steer, B.A., Schimmel, P. Genetic
code origins: tRNAs older than their synthetases? Proc. Natl. Acad. Sci. U.S.A.
Sardesai, N.Y., Schimmel, P. Noncovalent assembly of microhelix recognition
by a class II tRNA synthetase. J. Am. Chem. Soc. 120:3269, 1998.
Schimmel, P., Alexander, R. All you need is RNA. Science 281:658,
Schimmel, P., Alexander, R. Diverse RNA substrates for aminoacylation:
Clues to origins? Proc. Natl. Acad. Sci. U.S.A. 95:10351, 1998.
Schimmel, P., Söll, D. When protein engineering confronts the
tRNA world. Proc. Natl. Acad. Sci. U.S.A. 94:10007, 1997.
Schimmel, P., Tao, J., Hill, J. Aminoacyl tRNA synthetases as targets
for new anti-infectives. FASEB J., in press.
Sen, S., Zhou, H., Ripmaster, T., Hittleman, W.N., Schimmel, P., White,
R.A. Expression of a gene encoding a tRNA synthetase-like protein is enhanced
in tumorigenic human myeloid leukemia cells and is cell cycle stage- and differentiation-dependent.
Proc. Natl. Acad. Sci. U.S.A. 94:6164, 1997.
Shiba, K., Hiromi, H., Schimmel, P. Maintaining genetic code through
adaptations of tRNA synthetases to taxonomic domains. Trends Biochem. Sci. 22:453,
Shiba, K., Stello, T., Motegi, H., Noda, T., Musier-Forsyth, K., Schimmel,
P. Human lysyl-tRNA synthetase accepts N73 variants and rescues E coli double-defective
mutant. J. Biol. Chem. 272:22809, 1997.
Wakasugi, K., Quinn, C., Tao, N., Schimmel, P. Switching species-specific
aminoacylation with a peptide transplant. EMBO J. 17:297, 1998.
Whelihan, E.F., Schimmel, P. Rescuing an essential protein-RNA complex
with a nonessential appended domain. EMBO J. 16:2968, 1997.