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Methods in Molecular Biology; vol. 498. High Throughput Protein
Expression and Purification: Methods and Protocols, Sharon A. Doyle,
ed. Humana Press 2009, pp 322, ISBN: 978-1-58829-879-9
The end of the twentieth century brought
about two events which most significantly influenced the further development of molecular biology.
The first was the sequencing of several eukaryotic
genomes, including the human one. The second was
the discovery of the RNAi phenomenon. The former
had long been anticipated; the latter came as a surprise. Together, they drastically changed our thinking about the mechanisms controlling the release
and flow of genetic information from DNA through
RNA to proteins. In addition, they clearly showed
that scientists have reached a point at which further
progress in our understanding of complex biological systems requires that reductionist approaches be
replaced or complemented with more holistic ones.
Molecular biologists realized that the methods they
were using to study individual genes, transcripts, or
proteins are insufficient to investigate whole genomes, transcriptomes, or proteomes.
One of the major challenges facing modern
molecular biology is to give a detailed description
of cellular proteomes. Such a description requires
not only that all proteins occurring in a given cell
type be identified, but also that their biochemical
properties and their structures be determined. The
first stage on the long way to this goal is the elaboration of high throughput methods of protein production and purification. High Throughput Protein
Expression and Purification: Methods and Protocols, edited by Sharon A. Doyle, is an interesting attempt
showing what one currently has at one’s disposal
if one wishes to make a first step toward this aim.
The book is divided into twenty chapters whose
authors — seventy-two well-recognized specialists
— work at the best governmental laboratories and
private companies, all of them global leaders in
protein research.
The first chapter reviews modern strategies
of gene cloning, most effective prokaryotic and eukaryotic expression systems, and various methods of
protein purification and analysis. The second chapter shows how to design individual experiments and
find an optimal method of protein production. The
remaining eighteen (strictly methodological) chapters describe particular strategies or techniques that
have been successfully applied for high throughput
protein expression. Each of these chapters contains
a short summary and a concise theoretical introduction; it specifies all necessary materials and lists
numerous protocols which precisely explain how to
perform each experiment.
The methodological chapters can be further
divided into two groups. The first one provides readers with a wide spectrum of information on different
types of expression systems. Their key elements are
vectors into which one can insert any protein-coding
sequence. The authors have decided to present six
frequently used systems which, as they believe, are
especially well designed for high throughput cloning, clone screening and gene expression in E. coli,
yeast, insect and mammalian cells. By applying any
of these systems one should be able rapidly to select
clones capable of producing a target protein in high
amounts and in soluble form. Importantly, each of
the presented systems offers a unique cloning strategy. For example, the Gateway system is based on
recombinational cloning. Flexi vectors require ligation-dependent cloning and in the case of In-Fusion
and LIC vectors cloning is ligation-independent.
The second group of methodological chapters presents various approaches used to improve
the efficacy of protein production in prokaryotic,
eukaryotic or cell-free systems. In addition, they
show how applying specific protein tags, chromatographic techniques, detergents, or other chemicals can make it easier and faster to detect, isolate, and purify target proteins. Special attention
is paid to so-called “difficult proteins” which require extraordinary techniques and methods to be
expressed in sufficient amounts and proper form.
This problem concerns primarily highly hydrophobic membrane proteins.
Reading this review one can get the impression that the book edited by Doyle is yet another
handbook of protein expression and purification,
similar to many others published in the past years.
One crucial factor, however, singles it out from
among them. In the volume under review, the readers are shown how to change the scale of their experiment from one clone or protein to hundreds of
them. In addition, Doyle’s book helps the readers
learn how to apply the existing technologies, apparatus and robots to make automatic practically each
step of the protein production process: from cloning
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and clone screening, through protein expression and
purification, to the analysis of the final products. Accordingly, this book should be recommended especially to researchers who would like to organize a
new laboratory focused on high throughput protein
production, or to improve an existing one. It can
2009
also prove helpful to those who wish to optimize individual protein expression, isolation or purification.
Marek Figlerowicz
Institute of Bioorganic Chemistry
Polish Academy of Sciences
Noskowskiego 12/14,
61-704 Poznań, Poland
Methods in Molecular Biology vol. 517, Toll-Like Receptors, Methods
and Protocols, Claire E. McCoy and Luke A.J. O’Neill, eds. Series of
Springer Protocols. Humana Press, 2009, ISBN: 978-1-934115-72-5
At present immunology belongs to the most
dynamically developing biomedical sciences. Recent achievements in the field of innate immunity
dramatically have changed our knowledge of the
mechanisms involved in immune response. Discovery of Toll receptors in mid eighties ( awarded Nobel Prize in 1995) and further investigations resulted
with the recognition of Toll-like receptors (TLR), allowed to understand how immune cells recognize
various pathogen-associated molecules inducing of
both, innate and adaptive host defense response. To
date ten human TLRs and thirteen mouse TLRs have
been identified. TLRs contain leucine-reach repeats
(LRR) in their extra cellular domain and an intracellular region that share homology with interleukin1 receptor called TIR. Upon receptor activation the
TIR domain recruits downstream adaptor signaling
molecules i.e. protein kinases that activate transcription factors NFκB or IRFs and trigger specific genes
expression responsible for realization of the effector
function of the cell. The importance of TLRs in immune response and their implications in pathogenesis of various diseases are the subject of many investigations. Further understanding of the molecular
mechanisms of TLR signal transduction pathways,
involving receptor-ligand interactions and cascades
of intracellular signaling including posttranslational
modifications, is crucial to reveal its biological role
in health and disease. This is possible only by application and development of many analytical techniques and interdisciplinary studies combining new
achievements in the fields of molecular biology, genetics, biochemistry and immunology.
The book “Toll-Like Receptors: Methods and
Protocols” with 65 contributors, focuses on recent
developments of TLRs investigations performed
by many institutions including Universities, Technical Universities and Medical Schools in Europe
and USA. It comprise 25 chapters (447 pages) with
black- and- white illustrations and color plates, divided into four parts covering the different aspects
of TLRs studies.
The first part is focused on methods of detection and analysis of TLRs including analytical
RT-PCR approach for study of TLR expression at
the transcriptional level, expression of human TIR
domains in bacteria, purification and crystallization,
tools for in vitro studies of ligands and host cells
with the use of commercially available ligands, investigations of protein-protein interactions, subcellular localization and conformational changes in living
cells by confocal laser scanning microscopy (LSM)
with GFP technology, fluorescence resonance energy
transfer (FRET), bimolecular fluorescence complementation (BiFC) and combination of confocal imaging with FRET. This part also includes data on bioinformatic tools for analysis of TLR sequences and
three-dimensional structures, as well as for prediction and characterization of ligand binding. Experimental procedures for measurements of TLR interactions with their ligands are also described.
The second part consists chapters on methods
of analysis of the TLR down-stream signal transduction pathways. Key intermediates of the signaling
pathways are identified after expression in bacteria,
by liquid chromatography and tandem mass spectrometry (LC-SM/SM) or by differential in-gel electrophoresis DIGE (2D electrophoresis coupled with
protein labeling) using proteomic strategies. The
mammalian protein-protein interaction trap (MAPPIT) technique was used to study early signaling
steps from TLR4 to IRAK-1. The methodological approaches used to analyze the role of posttranslational modifications (phosphorylation, ubiquitination)
on TLR signaling pathways include transient transfections, site-directed mutagenesis, immunoprecipitation and immunobloting . Techniques for study of
TLR stimulation on apoptosis and negative regulation of TLR signaling, including influence of viral
proteins, are also described.
The third part describes genetic techniques
applied in TLR analysis. A practical guide is given
for performing germline mutagenesis in mouse using
ENU (N-ethyl-N-nitrosurea) for identification of the
genes involved in TLR signaling. Receptor functions
are investigated by microarray techniques using
Two-Color Microarray assay. Microarray processing and data analysis are described, including list of
software and web sites. Two very important new approaches are described. One is application of siRNA
Books reviews
Vol. 56 technology, a superior method of post-transcriptional gene silencing (PTGS) for uncovering novel gene
functions in TLR signaling. Second is the discovery
by genotypic methods of TLR polymorphism, that
affects host susceptibility towards infections, RLFP,
genotyping via the Sequenom and Illumina platforms and direct sequencing are p[resented.
The four part describes techniques applied to
study of the role of TLRs in various diseases with
use of experimental models of pathogen-host in vitro
and in vivo interactions. It involves investigations
of animal responsiveness to systemic challenge, isolated and cultured cell activation and proliferation
assays, reporter gene activation assay and immunohistochemistry (IHC) with immunofluorescence (IF).
The presented data include experimental disease
models of acute infection, septic shock and atherosclerosis, and in vitro investigations of cultured fibroblasts from rheumatoid arthritis patients and tissue samples of patients with collateral cancer. The
involvement of TLRs in adaptive immune response
is documented by studies of the activation of mouse
B cells and dendritic cells by immune complexes (IC)
characteristic of autoimmune diseases. The quantita-
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tive gene expression measurement assay based on
PCR technique with biomarkers of the TLR activity,
and its applications in clinical trials, presented also
in this part , will allow development of innovative
therapies.
All chapters provide short introduction, stepby- step laboratory protocols with lists of materials and reagents and, what is very important, their
sources. Experimental tips and troubleshooting procedures are also included, which is invaluable for
beginners in the field.
The covering of such wide range of problems
concerning investigations of the role of TLRs in the
innate and adaptive immune response makes the reviewed book an excellent manual for all investigators as well as teachers and advanced students of
biological and medical sciences interested in modern
immunology. That book fills an informational gap in
very vital field of knowledge.
Elżbieta Wałajtys-Rode
Department of Biochemistry and Biotechnology, Chemical Faculty,
Rzeszow University of Technology
Al. Powstańców Warszawy 6,
35-959 Rzeszów, Poland
Methods in Molecular Biology; vol. 541. Computational Systems
Biology, Jason McDermott, Ram Samudrala, Roger E. Bumbgarner,
Kristina Montgomery, Reneé Ireton, eds. Humana Press 2009.
The new volume of the Humana Press “Methods in Molecular Biology” series, entitled “Computational Systems Biology,” consists of 25 chapters
authored by 57 specialists in the field. As is noted in
one of these chapters (Chapter 20), a one-gene-onestudy approach is unlikely to enable the identification of the functions of all genes/products and understanding of a given cell or organism as a whole.
High-throughput approaches bring this goal closer,
and the huge amounts of data generated require
the development of new computational methods to
handle it and to use it to build useful models. This
book contains a wide collection of methods that can
be used with this goal in mind and gives a broad
review of this fascinating and quickly developing
field.
Part I of the book (“Network Components”)
starts with methods that are related to finding edges linking nodes in regulatory networks. Chapter
1 presents a quite detailed protocol of how to use
a specific software package (A-GLAM) to identify
cis-regulatory elements from the combination of coexpression data and functional annotations. Chapter 2 provides a different perspective: a method to
find transcription factor binding sites based on the
3-dimensional structures of transcription factors
complexed with DNA (the starting point structures
can be determined experimentally or by modelling).
Then, every base pair in DNA is changed to another
possible base pair. Molecular dynamics and free energy calculations are used to obtain the relative free
energies of binding to different sequences which can
then be combined to a single position weight matrix.
The next chapters are dedicated to the inference of
protein-protein interactions: using known interactions after a careful choice of protein domains that
explain the interactions (Chapter 3) or correlation of
evolutionary divergence (Chapter 4, with an important conclusion that the co-evolution model poorly
predicts protein-protein interactions). Chapter 5 is
more general and reviews algorithms used to integrate genome data to predict protein-protein interactions using machine learning. It gives some comments on the interaction data sets: for example, on
the bias towards important proteins and important
interactions in the positive interaction set and problems with constructing a negative data set (a set of
proteins that are known not to interact). The next
chapter (Chapter 6) is again a very detailed description of a method to use the information on transcriptional regulation in a model organism to infer regulatory interactions in a related organism. The procedure starts from finding similar gene pairs using sequence search. The predicted edges in the regulatory
network are then filtered by the search for binding
sites, functional annotation, protein localisation, etc.
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748
The last chapter in Part I (Chapter 7) stands out as
the only one that discusses metabolic networks. Not
only does it provide an exceptionally accessible introduction to the analysis of general network properties, but also describes a very interesting way to
reveal the hierarchy in biological networks. This is
also one of two chapters in the book accompanied
by colour plates.
The difference of focus between Part I and
Part II (“Network Inference”) of the book is hardly
noticeable. Chapter 8 considers again the problem
of inferring transcriptional regulatory networks in
non-model organisms. A simple approach is used:
if a transcription factor and its target gene have orthologs in the non-model genome, an interaction is
inferred. The authors consider significance of conservation of genes, interactions, network motifs (such
as feed-forward motif etc.) and global network structure. It may be slightly disappointing that results are
discussed only cursorily in Note 11 (p. 178). Chapter
9 provides a general introduction to the problem of
inference of transcriptional networks from data obtained using microarrays, protein-DNA and proteinprotein interaction assays, and functional annotations. It then proceeds to describe methods, giving
details on the authors’ own tool, Inferelator. The introduction to Chapter 10 starts with an observation
that differential gene expression is sometimes linked
to variation in specific genomic locations; methods
to find regulatory pathways from such expression
quantitative trait loci are first outlined, then evaluated using gene knockout data. The subject of regulatory network inference is continued in the last
chapter of Part II (Chapter 11), with emphasis on
Bayesian networks as an approach to obtaining networks with predictive value from expression data
and other data types.
In Part III (“Network Dynamics”), the focus
slowly shifts. Chapter 12 describes in intricate detail
two algorithms: CoExMiner, capable of handling not
only linear, but also nonlinear relationships of gene
co-expression and of suggesting directionality of
regulatory relationships, and PathwayPro, a tool to
simulate the effects of perturbations to the network
(such as a change of one or several gene expressions) on its complex behaviour, as a way to identify targets for therapeutic intervention. Chapter 13 is
even more mathematically advanced and describes
the theory to analyse the dynamics of biochemical networks (biochemical control theory) with applications to some basic motifs in cellular networks
(e.g. branches or feedback loops), with important
conclusions for those wishing to target pathways.
The chapter ends with a comprehensive reading
list. While Chapter 13 concentrates on deterministic
models, Chapter 14 is somewhat complementary, as
it introduces the approaches to address stochastic
2009
fluctuations in the concentration of biomolecules. It
provides an extensive list of software tools for kinetic simulations of systems. The last chapter of Part III
(Chapter 15) would be of interest to molecular biologists that might wish to apply their skills to providing the computational biologists with high quality
data. As an example, the authors describe their own
study of promoter regulation in phage λ in which
computational modelling allowed them to measure
the energetics of DNA looping.
Part IV (“Function and Evolutionary Systems
Biology”) starts with a chapter presenting a model
of intron evolution (Chapter 16). The authors derive
very interesting conclusions from their analysis (p.
360), for example, that the frequencies of intron gain
and loss are correlated (suggesting a common mechanism for both processes) and that the frequency of
intron gain is correlated with gene expression level.
Sadly, the results themselves are not presented; the
bulk of the chapter is dedicated to a very detailed
analysis of the algorithm. Chapter 17 is dedicated to
functional prediction (with an example of prediction
of the EC number and Pfam classification) from features extracted from sequences (such as amino acid
composition, average hydrophobicity, protein length
etc.) and data such as tissue specificity, subcellular
localisation, cofactors etc. The authors’ algorithm
uses mixtures of stochastic decision trees with novel criterion functions to evaluate tree performance.
The next chapter (Chapter 18) presents a method to
identify specificity-determining residues (in brief,
conserved in orthologs but different in paralogs)
and residues co-evolving in interacting proteins. An
example of identifying such residues in bacterial
two-component (histidine kinase/response protein)
systems is provided. Colour plates are used to illustrate the results. Chapter 19 describes how to take
advantage of the information about protein-protein
interactions to understand clinical phenotypes: genes
coding for proteins interacting with known disease-associated proteins are better candidates when
searching for disease-causing mutations among
polymorphisms. A method to discriminate between
damaging and neutral mutations using support vector machines is presented. Chapter 20 discusses the
problem of functional bias in gene annotations. Such
bias can affect further discovery of gene functions
from expression data. For example, out of more than
1000 functional annotation terms, one (‘protein biosynthesis’ ) covers more than 4% of annotations. The
authors observed that removing proteins pairs sharing this term resulted in lower correlation between
the likelihood of functional coupling and co-expression, while the correlation between the likelihood of
functional coupling and the measure of protein-protein interaction remained unaffected. They propose
that masking dominant terms can help when ma-
Books reviews
Vol. 56 chine learning methods are used to infer interactions
from gene expression data.
Part V (“Computational Infrastructure for
Systems Biology”), the last part of the book, starts
with a chapter on methods to assess algorithms for
clustering gene expression data (Chapter 21). After
a brief introduction to several clustering algorithms,
the authors consider the problem of determining
whether the structure revealed by such algorithms is
meaningful and if it did not occur by chance or as
an artifact of the algorithm itself. Two complementary indices are introduced, one based on the minimal
description length principle, the other using experimentally verified data sets to assess the quality of
clustering. Chapters 22 and 23 are dedicated to Bioverse, a framework for handling biological information based on hierarchical levels of organization, developed by the editors of the book. The final chapter
(Chapter 24) is devoted to Software Environment for
BIological Network Inference (SEBINI) and a tool
for Collective Analysis of Biological Interaction Networks (CABIN). SEBINI is an open-source platform
which allows (among other things) the use of many
inference methods (based on correlation, Bayesian
networks, information theory) and algorithm comparison. The networks exported from SEBINI can be
integrated with other data and further analysed using CABIN.
All in all, “Computational Systems Biology”
is a rather uneven volume, with some chapters providing a general, original perspective and others that
are rather mundane, with perhaps unnecessarily de-
749
tailed lists of hardware requirements, programme
running options, etc., expected rather in software
documentation. On the other hand, when experiments or results are presented, in some chapters it is
done perhaps too sketchily and the reader may need
to search for the original papers by the authors of a
given chapter. A quick glimpse at some of the equations reveals problems with notation (e.g., p. 45) or
editing errors (e.g., pp. 75, 193, and 493), which may
worry someone wishing to follow more carefully the
derivations in chapters with are heavy with lengthy
equations (such as Chapter 16, with over 30).
As mentioned by one of the book authors,
Herbert M. Sauro (p. 270), molecular biology traditionally did not have much need for mathematics.
Now with the need to handle enormous amounts of
data that are being generated by high-throughput
experiments, biology becomes more and more interesting for those inclined rather to deal with equations and algorithms than with small volumes of liquids. “Computational Systems Biology” is definitely
not an introductory text, and it will not be of much
help to a wet-lab molecular biologist or a biology
undergraduate that wishes to get a glimpse of the
field. But for a computational biologist, it is a fascinating read, a broad and comprehensive resource on
the current methods and approaches.
Borys Wróbel
Computational Biology Group,
Institute of Oceanology,
Polish Academy of Sciences,
Powstańców Warszawy 55,
81-712 Sopot, Poland
Plant Genomics. Methods and Protocols, Somers DJ, Langridge P,
Gustafson JP, eds. Humana Press 2009, pp 362, ISBN: 978-1-58829-997-0
The book is latest volume in the series “Methods in Molecular Biology” under a general editorial
overview by JM Walker. The main aim of the series
is to present research protocols for golden standards
of molecular biology methods.
The volume “Plant Genomics. Methods and
Protocols” concerns a wide spectrum of problems,
some of them outside of the main scope of the field,
e.g. transcriptomics and proteomics. The editors, DJ
Sommers, P Langridge and JP Gustafson, invited 43
contributors from the United States, United Kingdom, Australia, Canada, India, Israel and Germany
to present modern methods covering the interests
of those working in the diverse fields of plant molecular: biology, ecophysiology and breeding as well
as biotechnology and bioinformatics. Some of the
methods will also be of interest to readers working
in plant cell biology and plant physiology.
The book is divided into 18 chapters of three
types. First, some chapters concentrate on wet work
and are strictly oriented on the practical aspects of a
given method, including short introduction, step-bystep protocols, a list of materials and chemicals, tips
and tricks, and troubleshooting notes. Of the second
type are chapters on bioinformatics, containing descriptions of different data bases and principles of
working with them. The remaining chapters present
either sort of a review on genomic technologies or
description of complete scientific projects.
The first chapter of the volume describes the
role of model plant species in advancing our knowledge of plant functioning at the molecular level over
the past 25 years. It is followed by several chapters
focusing on molecular genetic markers. These chapters concern ultra-high throughput genotyping, construction of genetic linkage maps, and generation of
large-insert BAC libraries. Also, description of a new
method of identifying genetic markers by means of
analysis of bundance from cRNA hybridizations to
Affymetrix microarrays is presented.
Another set of chapters concerns functional
genomics. First, overviews of both stable and tran-
Books reviews
750
sient transformations are presented. Forward and
reverse genetic strategies are described for assigning
functions to newly identified genes. The in situ hybridization method is presented as a powerful tool
for determining the spatial and temporal expression
patterns of a given gene. Such information is important for determining the cellular and/or tissue as
well as developmental stage specifity of gene action.
Methods used in proteomics as well as heterologous
and cell-free protein expression systems are also described, as they allow more accurate, experimental
assignment of gene functions. Also, techniques used
is structural biology are reviewed, aimed at the determination of molecular structures at the three-dimensional (3D) level, which, in turn, is important
for understanding enzyme functions, protein-protein
interactions and transcription factor-DNA interactions.
Nowadays, databases, bioinformatics and
computational biology have been integrated into
biological research. This is reflected in two chapters
of the volume. The first one concentrates on major
open-access plant and crop databases. The second
chapter is focused on genome annotation methods,
including sequence quality, structural annotation,
functional annotation and visualization tools. It includes an overview of the basic sources of genomic
sequences along with methods and resources of use
to researchers in the structural and functional annotation of gene sequences. Some examples of application of selected tools are presented.
Four last chapters concern more general aspects of plant genomics by presenting examples of
either basic or applied research projects. The chapter “Molecular Plant Breeding: Methodology and
Achievements” shows several examples of success-
2009
ful use of of molecular breeding in cereals. It also
describe step-by-step procedure of a marker-assisted
breeding project. In another chapter, “Practical Delivery of Genes to the Marketplace”, practical application of gene discovery is described. Steps necessary
before commercializing a crop with a biotechnology
trait are described (including the regulatory process
in the USA, with reference to other countries, excluding, however, the European Union). The chapter “Ecological Genomics of Plant Populations: The
Israeli Perspective” describes a research program on
ecological genomics of wild progenitors of wheat
and barley. In the project, hundreds of populations
and thousands of genotypes have been analyzed for
protein and DNA sequence diversity, and responses
to abiotic and biotic stresses analyzed. Additionally, genetic maps, cloning and transformation of
candidate genes were introduced to the project. The
last chapter of the volume is entitled “Genome Sequencing Approaches and Successes”. It describes
sequencing technologies, including the current, a
low-cost ones developed by Roche, Illumina/Solexa,
and Applied Biosystems. Examples of plant genome
sequencing projects are also presented thus elegantly
closing the frame opened by the Introductory Chapter 1 entitled “Role of Model Plant Species”.
In summary, “Plant Genomics. Methods and
Protocols” can be recommended as a valuable source
of timely information for students, researchers and
breeders.
Paweł Sowiński
Department of Plant Molecular Ecophysiology
Institute of Plant Experimental Biology
Faculty of Biology, University of Warsaw
Miecznikowa 1,
02-096 Warszawa, Poland
Essentials of Apoptosis, A guide for basic and clinical research, XiaoMing Yan and Zheng Dong, eds. Humana Press, 2009, 2nd edn.
The programmed cell death called apoptosis is
a universal phenomenon in the living world. When a
cell is severely damaged, for example is infected by
a virus, or its DNA is damaged beyond repair – the
apoptotic cascade of molecular events may eliminate
such a cell from the organism. Moreover, in many
cases apoptosis is a part of a physiological process,
where cell death is required for the proper development of tissues and organs. Cell death has been described already in 1842 by the German scientist Carl
Vogt. The last two decades experienced a plethora
of data on the molecular mechanisms of apoptosis,
and the database PubMed shows over 175 000 papers which deal with the process of programmed
cell death.
The book “Essentials of Apoptosis” is a collection of 31 review articles, covering a wide variety
of approaches in studying cell death. Part one covers
the molecules and pathways of apoptosis, part two
presents studies of apoptosis in model organisms:
plants, yeast, C. elegans and Drosophila, part three
discusses apoptosis in mammalian physiology and
pathogenesis, and finally part four concerns alternative cell death mechanisms: necrosis, autophagy,
caspase-independent cell death and cell death depending on lysosomal proteases.
This wide scope of the reviews included in
the book constitutes its major value. Written by experts in their respective fields of research, each article presents a summary of up-to date knowledge,
allowing the reader to get an insight into molecular
mechanisms of cell death.
While I feel the book is a highly recommended source of reference information to any researcher
Books reviews
Vol. 56 interested in apoptosis, it has some minor flaws. To
my mind most articles would benefit from including
of more schematic representations of the pathways
discussed. In some cases the text describes a large
collection of proteins and complexes participating in
a given pathway, with no or very incomplete graphic representation or. Too long paragraphs heavily
loaded with gene and protein names occasionally
make the text poorly difficult to follow. In some cases a stronger control from the editors would seem
advisable, both in suggesting the authors to make
themselves more clear, and in removing “cloned”
fragments such as exact repetition of the summary
as the introduction.
751
The critical remarks listed above do not diminish my enthusiasm about this book. It enables
the reader to get acquainted with many aspects of
apoptosis, and impressive amounts of references
cited in each of the article allows for further studies
on the subject. I recommend this book to anyone
seriously interested in research on programmed cell
death.
Piotr P. Stepien
Institute of Genetics and Biotechnology
University of Warsaw,
Pawińskiego 5a,
02-106 Warszawa, Poland
Handbook of Modern Biophysics, vol. 1, Fundamental Concepts in
Biophysics, Thomas Jue ed. Humana Press, 2009, pp 240,
ISBN: 978-1-58829-973-4
Thomas Jue from the University of California
Davis, has introduced a new series of books under
the title Handbook of Modern Biophysics. As we read
in the preface „The books in this series will bring
current biophysics topics into focus and expand as
the field of biophysics expands, so that biology and
physical–science students or researchers can learn
fundamental concepts and apply new biophysics
techniques to address biomedical questions.” We
have just received the first volume Fundamental Concepts in Biophysics. It contains seven chapters written by 13 contributors, all form the University of
California Davis. Each chapter refers to a particular
problem from the fields of theoretical or mathematical and experimental physics underlying biophysical
experiments and their interpretation. Each chapter
concludes with a set of problems which help the
reader to understand and further analyse the ideas
presented. The chapters contain also lists of further
reading and references.
In the first chapter Mathematical Methods in
Biophysics Rajiv R.P. Singh presents some basic concepts from the probability theory. It is assumed to
be a useful introduction to the diffusion equation,
which is discussed further, and a general solution of
this equation is presented.
The second chapter Quantum Mechanics Basic
to Biophysical Methods by William Fink starts with a
very brief presentation of the postulates of quantum
mechanics together with the introduction of the operator and its eigenvalue problem, a short description of Heisenberg’s and Schrödinger’s formulation
of quantum mechanics and the Born interpretation
of this theory. These basic concepts are than applied in the description of a one-dimensional motion
of a particle, the model of the harmonic oscillator,
and the hydrogen atom. Since exact solutions of the
Schrödinger equation are known only for a few simple problems, the author presents shortly two fundamental approaches which lead to approximate
solutions: perturbation theory and the variational
principle.
Numerous crucial cellular processes are related to the phenomenon of receptor — ligand binding. The most important is cellular signalling. Our
current knowledge about receptor — ligand binding
and about molecular recognition in general, is based
not only on experimental data but also on the results
of computational modelling. Two classes of models
are used in these calculations: differential equationsbased models and Monte Carlo models. Application
of any of these models depends on the assumptions
about molecular concentrations and the spatial homogeneity of their distribution. Both types of modelling are described in the third chapter written by
S. Raychaudhuri, Ph. Tsourkas and E. Willgohs.
The authors discuss application of modelling in the
analysis of enzyme reactions and the interaction of
T cells with antigen presenting cells. This chapter
completes the theoretical part of the book.
The next four chapters present a choice of
experimental methods used in biophysics. This
part opens with the paper on Fluorescence Spectroscopy by Y. Yeh, S. Fore, and H. Wu. After a short
introduction to the phenomenon of fluorescence the
authors describe modern techniques in fluorescence
microscopy together with several types of biological
fluorophores, like molecular fluorophores designed
for biological studies, a family of green fluorescent
protein (GFP), and quantum dots. This chapter contains also a brief description of several methods of
fluorescence spectroscopy, like fluorescence lifetime
spectroscopy, fluorescence anisotropy, photobleaching and fluorescence recovery after photobleach-
Books reviews
752
ing (FRAP), fluorescence resonance energy transfer
(FRET), and fluorescence correlation spectroscopy
(FCS).
The subsequent chapter by T-Y. Chen, Y-F. Lin
and J. Zheng contains an analysis of membrane bioelectricity. It starts with the presentation of the two
factors that drive ion movements through the membrane. One of them is the electrical driving force,
which originates from the difference in voltages in
the intracellular and extracellular sides. The second
factor is named chemical driving force. It results
from unequal ion concentrations on the two sides of
the biological membrane. The methods of voltage–
clamp and patch–clamp are applied in the analysis
of membrane bioelectricity. The two-electrode voltage-clamp method (TEVC) could be applied, e.g., in
the analysis of Xenopus oocytes. Patch-clamp techniques are applied in the analysis of individual ion
channels. A patch-clamp technique can be combined
with selected methods of fluorescence spectroscopy,
which enable one to analyse not only the flux of ions
but also the structural changes within a channel.
Single-particle techniques are the most important modern methods among those applied in
the studies of molecular and cellular biology. These
techniques are presented in the chapter Single – Particle Tracking by M.J. Saxton. Particles with a wide
variety of chemical and physical properties are used,
like small dye molecules, fluorescent proteins, quantum dots, latex or gold beads. Advantages and disadvantages of these are presented. The author discusses a few examples of the application of single–
particle analysis, among others in the analysis of the
infection of a cell by a virus.
The final chapter by Thomas Jue concerns the
application of NMR techniques in the analysis of
molecular diffusion. Two procedures are in use. The
2009
relaxation approach suffers from a number of limitations connected mainly with the requirement for
definition of the relaxation mechanism. The second
approach based on pulse-field gradient technique
(PFG) avoids these limitations by measuring the
spin–echo signal intensity as a function of the applied field gradient. Analysis of the signal attenuation caused by molecular self-diffusion enables one
to determine the translational diffusion coefficient.
The author describes the basic principles of the PFG
technique and presents its application in studies of
myoglobin-facilitated diffusion of oxygen in muscle
tissue.
Reading of the first volume of the series
Handbook of Modern Biophysics leads to the conclusion
that this form of presentation of modern biophysical
problems is very useful from the educational point
of view. It enables thorough presentation of a particular experimental technique. The chapters concerning the single-particle methods and NMR studies
are the best examples of this approach. There are no
fixed limits for the length of the contribution, as it is
usually observed in a textbook on biophysics. This
form of presentation could be also very useful for
the presentation of theoretical problems. For example the analysis of the diffusion equation is very important and very helpful, although it does not need
to start with the basic ideas of probability theory.
The series on modern biophysics should be
very helpful for students and young scientists working in molecular biology, biochemistry or molecular
physics.
Genowefa Ślósarek
Department of Molecular Biophysics,
Faculty of Physics
Adam Mickiewicz University,
Umultowska 85,
61-614 Poznań, Poland
G Protein-Coupled Receptors in Drug Discovery, Wayne R. Leifert ed.
Springer Protocols, Methods and Protocols, Humana Press, 2009,
ISBN 978-1-60327-316-9
G Protein-Coupled Receptors (GPCRs) form
a critical part of the protein repertoire used by
cells to sense their environment. Due to their mediation of numerous critical physiological functions, GPCRs are involved in most disease areas. It
is estimated that 40-50% of the current drugs target GPCRs. To fully understand the GPCR drug
discovery process one should study not only the
biology/chemistry of GPCRs but also have wide
knowledge of the methods/protocols used in this
process. This book serves this exact purpose.
The reviewed book consists of 26 chapters written
by scientists working with GPCRs either in pharmaceutical companies or top world universities. While
the first five chapters give an overview of GPCR
function, signalling and structural biology, the book
requires and assumes a certain level of knowledge
of GPCRs and their biology/chemistry. Most of the
chapters have only short introduction parts and
concentrate on giving a detailed, step-by-step description of protocols with notes on possible pitfalls.
The very interesting chapter 7 gives description the
homology modelling of GPCRs. This computational
method is the most commonly used approach to obtain high-quality GPCR models based on experimental structures determined using X-ray technology.
Authors of this part lead the reader through all steps
needed to obtain a computational model and succeed in explaining the whole process is simple terms.
Since computational tools are becoming more and
Books reviews
Vol. 56 more important in drug discovery, all experimental
scientists involved in this process should familiarize
themselves with this part to get ideas about basic theoretical methods available in their field of research.
Most of the book is, however, devoted to various
GPCR-connected assays. This part starts with Chapter 6, which describes the quality assessment of the
assay data. It is followed by Chapter 8 explaining
one possible way of GPCR expression. The next
eight chapters provide example of different strategies of GPCR-bases assays, which are raging from
radioligand and fluorescent binding assays to gene
delivery-based and solid substrate assays. The assays are described in details and many examples
which makes it easy to reproduce them. They are
also followed by detailed references, useful for
further exploration of the nuances of each technique and giving even more examples of their use.
Final chapters of this book are introducing some
of the latest, cutting-edge methods in drug discovery. Chapters 18, 20 and 21 are devoted to fluorescence-based techniques and their applicability to
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measurements of GPCR conformational changes
and ligand-receptor orientation. Chapters 22, 23
and 26 describe novel protocols for detecting GPCRs activation and their interactions with other
proteins. Finally, chapter 25 illustrates oocyte electrophysiology method used to deorphanize GPCRs.
Overall, this is a very interesting book that should
appeal to any scientists working in the GPCR drug
discovery field. It’s biggest strength is a clear description of every aspect of GPCR drug discovery
process, which is easy to understand even for a nonexpert in the respective field. This book is a mustread for everyone involved in GPCR drug discovery
and particularly for scientists working in multidisciplinary teams, who need to soundly communicate
ideas with experts in other fields.
Bartosz Trzaskowski
Laboratory of Biomodelling
International Institute of Molecular and Cell
Biology in Warsaw
Ks. Trojdena 4,
02-109 Warszawa, Poland