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Engineering the Genetic Code. Expanding the Amino Acid Repertoire for the Design of Novel Proteins. By Nediljko Budisa

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Engineering the Genetic Code
Expanding the
Amino Acid Repertoire for the Design
of Novel Proteins.
By Nediljko Budisa.
Wiley-VCH, Weinheim 2006.
296 pp., hardcover
E 119.00.—ISBN
Site-specific mutagenesis, a procedure
for replacing amino acids in proteins,
has improved since the late 1970s, when
it involved time-consuming experiments, to become a well-established
method with ready-to-go kits. It is now
standard practice to modify proteins in
this way, and examples range from the
analysis of the mechanistic basis of
enzyme reactions to the optimization
of proteins for biotechnological use.
This calls for more than the site-specific
introduction of the canonical amino
acids, the 20 proteinogenic ones contained in the universal genetic code.
Experiments to introduce non-canonical
amino acids into the proteome were
already being carried out in the 1950s. In
continuation of work by Cowie and
Cohen, the replacement of methionine
by selenomethionine is regularly used
today. So, how far have we come after
50 years? This book describes the capabilities of modern “genetic code engineering”, which refers to methods for
producing proteins (alloproteins) with
new peptide building blocks by ribosomal protein biosynthesis.
To put together a textbook in such a
topical and fast-moving field within a
short enough time is often only possible
by deciding on a multi-author work
Angew. Chem. Int. Ed. 2006, 45, 3903
managed by an editor. Fortunately,
Nediljko Budisa has taken on the task
of writing about the complex field in one
go. Budisa has collected examples of the
incorporation of more than 150 new
amino acids, mostly in overexpressed
proteins. He describes at length the
necessary background of the translation
machinery, the universal genetic code
and the amino acids that are represented,
the concepts of genetic code engineering
with examples, the evolution and malleability of the genetic code, and applications of genetic code engineering.
The first two chapters give definitions of technical terms and a historical
overview. New terms and their definitions are necessary in an emerging field.
Some of them are given at the beginning, others are distributed throughout
the book. A glossary would add readability and offer the chance to develop a
Chapter 3
describes all the steps needed to proceed
from having a free amino acid in the
cytosol to inserting it at its destination in
the polypeptide chain. This book is not a
textbook of biochemistry, and therefore
the reader needs to know the basics of
translation, and is expected to know
about established methods in molecular
biology and protein chemistry. For university teaching, the book offers material for an advanced lecture course; for
the scientist at the bench it is an up-todate compendium that will enable him
to add genetic code engineering to his
At the core of the book are the
concepts and methods of genetic code
engineering, and descriptions of how
they have been applied to homologues
and more distant derivatives of several
of the proteinogenic amino acids (Chapter 5). The range of methods described
includes simply feeding the selected
amino acids to auxotrophic organisms,
modifying the specificity of the tRNA or
(AARS) or amino acid metabolism,
and even the construction of a complete
system (in this case an Escherichia coli
strain) with matched metabolism,
tRNA, and AARS. In almost all cases,
synthetically prepared amino acids must
be added to the medium and are
accepted by the ribosomal protein biosynthesis. In contrast, in the most
advanced method the metabolism sup-
plies the desired amino acid, and then
the orthogonal AARS charges the additional specific tRNA. With help of this
tRNA, the new amino acid can be
introduced at the defined positions, for
example by suppression of a UAG stop
codon. Although this is an elegant
method for alloprotein synthesis, one
must not forget the immense effort that
this requires in the laboratory. Only the
group of P. G. Schulz has so far succeeded in constructing such an artificial
E. coli; their strain incorporates p-aminophenylalanine into proteins.
The last chapter discusses expectations from genetic code engineering
and describes some successful applications. By using fluorinated amino acids it
may be possible to prepare fluorophilic
proteins with neither hydrophilic nor
hydrophobic properties. It is hoped that
by this means one can achieve protein
stability in aqueous and organic solvents. An example of a successful application is the replacement of the chromophore-forming tyrosine (Tyr66) of
green fluorescent protein (GFP) by
other amino acids with modified aromatic side chains. The author<s competence and long-term involvement in this
field is visible in the thoroughness with
which he has collected examples. However, in some places, for example in the
description of posttranslational modifications, a table might well have supplemented or replaced some text.
The numerous examples illustrate
the new prospects that are opened up by
genetic code engineering. We can expect
novel proteins with properties that
cannot be foreseen in detail, when one
considers that, even for polypeptides
based on the 20 proteinogenic amino
acids, the rules of folding are still not
known. Additional topics are new applications in technology and pharmacology, evolution of the genetic code, and
the ethics of the far-reaching changes
that genetic code engineering makes
possible. Budisa rates the environmental
risk potential of new applications as low
compared to the expected benefits, thus
launching a necessary discussion.
Gottfried J. Palm
Institute for Chemistry and Biochemistry
University of Greifswald (Germany)
DOI: 10.1002/anie.200585385
4 2006 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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acid, engineering, design, expanding, repertoire, amin, protein, novem, genetics, nediljko, code, budisa
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