KEY CONCEPTS:
- Aminoacyl-tRNA synthetases are divided into the class I and class II groups by sequence and structural similarities.
Synthetases have been divided into two general groups, each
containing 10 enzymes, on the basis of the structure of the domain that contains
the active site. A general type of organization that applies to both groups is
represented in Figure 7.14. The catalytic domain includes
the binding sites for ATP and amino acid. It can be recognized as a large region
that is interrupted by an insertion of the domain that binds the acceptor helix
of the tRNA. This places the terminus of the tRNA in proximity to the catalytic
site. A separate domain binds the anticodon region of tRNA. Those synthetases
that are multimeric also possess an oligomerization domain (for review see Schimmel, 1987).
Class I synthetases have an N-terminal catalytic domain that
is identified by the presence of two short, partly conserved sequences of amino
acids, sometimes called "signature sequences." The catalytic domain takes the
form of a motif called a nucleotide-binding fold (which is also found in other
classes of enzymes that bind nucleotides). The nucleotide fold consists of
alternating parallel β-strands and α-helices; the signature sequence forms part of the
ATP-binding site. The insertion that contacts the acceptor helix of tRNA differs
widely between different class I enzymes. The C-terminal domains of the class I
synthetases, which include the tRNA anticodon-binding domain and any
oligomerization domain, also are quite different from one another.
Class II enzymes share three rather general similarities of
sequence in their catalytic domains. The active site contains a large
antiparallel β-sheet surrounded by α-helices. Again, the acceptor helix-binding domain that
interrupts the catalytic domain has a structure that depends on the individual
enzyme. The anticodon-binding domain tends to be N-terminal. The location of any
oligomerization domain is widely variable.
The lack of any apparent relationship between the two groups
of synthetases is a puzzle. Perhaps they evolved independently of one another.
This makes it seem possible even that an early form of life could have existed
with proteins that were made up of just the 10 amino acids coded by one type or
the other.
A general model for synthetase·tRNA binding suggests that the protein binds the tRNA
along the "side" of the L-shaped molecule. The same general principle applies
for all synthetase·tRNA binding: the tRNA is bound
principally at its two extremities, and most of the tRNA sequence is not
involved in recognition by a synthetase. However, the detailed nature of the
interaction is different between class I and class II enzymes, as can be seen
from the models of Figure 7.15, which are based on
crystal structures. The two types of enzyme approach the tRNA from opposite
sides, with the result that the tRNA-protein models look almost like mirror
images of one another.
A class I enzyme (Gln-tRNA synthetase) approaches the D-loop
side of the tRNA. It recognizes the minor groove of the acceptor stem at one end
of the binding site, and interacts with the anticodon loop at the other end. Figure 7.16 is a diagrammatic representation of the crystal
structure of the tRNAGln·synthetase
complex. A revealing feature of the structure is that contacts with the enzyme
change the structure of the tRNA at two important points. These can be seen by
comparing the dotted and solid lines in the anticodon loop and acceptor
stem:
- Bases U35 and U36 in the anticodon loop are pulled farther out of the tRNA into the protein.
- The end of the acceptor stem is seriously distorted, with the result that base pairing between U1 and A72 is disrupted. The single-stranded end of the stem pokes into a deep pocket in the synthetase protein, which also contains the binding site for ATP.
This structure explains why changes in U35, G73, or the
U1-A72 base pair affect the recognition of the tRNA by its synthetase. At all of
these positions, hydrogen bonding occurs between the protein and tRNA (Rould et al., 1989).
A class II enzyme (Asp-tRNA synthetase) approaches the tRNA
from the other side, and recognizes the variable loop, and the major groove of
the acceptor stem, as drawn in Figure 7.17. The acceptor
stem remains in its regular helical conformation. ATP is probably bound near to
the terminal adenine. At the other end of the binding site, there is a tight
contact with the anticodon loop, which has a change in conformation that allows
the anticodon to be in close contact with the protein (Ruff et al., 1991).