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chapter 25
RNA and Protein Synthesis
the 5' terminus itself of a mitochondrial mRNA molecule
binds to the ribosome. The plethora of diseases and clini-
cal abnormalities caused by mitochondrial mutations dis-
cussed in Chapter 14.
25.7 Attachment of Amino Acid
to tRNA Molecule
When an amino acid is covalently linked to a tRNA
molecule, the tRNA is said to be
aminoacylated
or
charged.
The notation for a tRNA charged with serine is
seryl tRNA. The term
uncharged tRNA
refers to a tRNA
molecule lacking an amino acid.
Acylation is accomplished in two steps, both of which
are catalyzed by an
aminoacyl tRNA synthetase.
In the
first, or activation, step an aminoacyl AMP is generated in
a reaction between an amino acid and ATP:
h
o
h
o
o
I
II
I
II
I
H3N +— C — C — O " + ATP
H3N +— C — C— O — P— Ribose— Adenine + PP
I
I
I
R
R
0 ‘
In the second, or transfer, step, the aminoacyl AMP reacts
with tRNA to form an aminoacylated tRNA and AMP:
Aminoacyl AMP + tRNA ^ aminoacyl tRNA + AMP
As in DNA synthesis, the reaction is driven to the right by
hydrolysis of pyrophosphate, so the overall reaction is
Amino acid + ATP + tRNA + H
2
O ->
aminoacyl tRNA + AMP + 2
Pj
Usually only one aminoacyl synthetase exists for each
amino acid. However, for a few amino acids specified
by more than one codon, more than one synthetase does
exist.
25.8 Initiator tRNA Molecules and Selection of
Initiation Codon
The initiator tRNA molecule in prokaryotes—tRNA™61—
has several properties that distinguish it from all other
tRNA molecules. One feature is that the tRNA is first
acylated with methionine, and then the methionine is
modified. Acylation is by methionyl tRNA synthetase,
which also charges tRNAMet. However, the methionine
of charged tRNA™61 is immediately recognized by an-
other enzyme, tRNA methionyl transformylase, which
transfers a formyl group from N10-formyltetrahydrofolate
(fTHF) to the amino group of the methionine to form
h
o
tRNA,M” + Met
, H3N — C— C — O — tRNA,M<’'
0
H
H
0
w
1
1
II
f
1
Z
1
- O
1
O
/
H
(CH2)2
1
1
s
j
CH3
— O — tRNA,MW
Transformylase
7
^
(CH3)2
I
s
I
♦C H3
N-formylmethionine (fMet): Transformylase does not
formylate methionyl tRNAMet
because tRNAMet and
tRNA™61 are structurally different. A second feature is
its ability to initiate polypeptide synthesis.
Eukaryotic initiator tRNA molecules differ from the
prokaryotic initiator molecule in several ways. The most
striking difference is that whereas eukaryotic organisms
produce both a normal tRNAMet and an initiator tRNA,
which is also charged with methionine, the methionine
does not undergo formylation. In eukaryotes, the first
amino acid in a growing polypeptide chain is Met and
not fMet. The codon for both kinds of tRNA molecules in
eukaryotes is AUG, just as for prokaryotes.
The sequence AUG serves as both an initiation codon
and the codon for methionine. Since methionine occurs
within protein chains, some signal in the base sequence of
the mRNA must identify particular AUG triplets as start
codons. In eukaryotes, initiation usually begins at the AUG
triplet nearest to the 5' terminus of the mRNA molecule,
i.e., no particular signal is used (although sequences do
play a role in the efficiency of initiation). Presumably, the
ribosome and the mRNA interact at the 5' terminus and
slide with respect to one another until an AUG is encoun-
tered. This AUG determines the reading frame. When a
stop codon is encountered, the ribosome and the mRNA
dissociate. Only a unique polypeptide is translated from a
particular eukaryotic mRNA molecule.
In prokaryotes, the situation is quite different. At a fixed
distance upstream from each AUG codon used for initia-
tion is the sequence AGGAGGU (ora single-base variant).
This sequence, known as the
Shine-Dalgarno sequence,
is complementary to a portion of the 3'-terminal sequence
of one of the RNA molecules in the ribosome (in partic-
ular, the 16S rRNA in the 30S particle; see below). Base
pairing between these complementary sequences orients
the initiating AUG codon on the ribosome. Thus, initiation
in prokaryotes is restricted to an AUG on the 3' side of the
Shine-Dalgarno sequence.
Many prokaryotic mRNA molecules are polycistronic
and contain coding sequences for several polypeptides.
Thus, a polycistronic mRNA molecule must possess a
