Protein Folding Protocols

343 Pages·2007·5.83 MB·English

METHODS IN MOLECULAR BIOLOGY™ 350
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Protein Folding Protocols
M E T H O D S I N M O L E C U L A R B I O L O G Y™
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M E T H O D S I N M O L E C U L A R B I O L O G Y™
Protein Folding
Protocols
Edited by
Yawen Bai
Laboratory of Biochemistry, National Cancer Institute,
National Institutes of Health, Bethesda, MD
and
Ruth Nussinov
Center for Cancer Research Nanobiology Program SAIC,
National Cancer Institute-Frederick, Frederick, MD
and Department of Human Genetics, Sackler School of Medicine,
Tel Aviv University, Tel Aviv, Israel
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Protein folding protocols / edited by Yawen Bai and Ruth Nussinov.
p. ; cm. -- (Methods in molecular biology ; 350)
Includes bibliographical references and index.
ISBN 1-58829-622-9 (alk. paper)
1. Protein folding. 2. Proteins--Conformation. 3. Proteins--Analysis.
[DNLM: 1. Protein Folding. 2. Protein Conformation. 3. Proteins--analysis. QU 55 P96635 2006] I. Bai,
Yawen. II. Nussinov, Ruth. III. Series: Methods in molecular biology (Clifton, N.J.) ; 350.
QP551.P695822 2006
572'.633--dc22
2006002791
Preface
Protein Folding Protocols presents protocols for studying and characterizing
steps and conformational ensembles populating pathways in protein folding
from the unfolded to the folded state. It further presents a sample of approaches
toward the prediction of protein structure starting from the amino acid sequence,
in the absence of overall homologous sequences. Protein folding is a crucial
step in the transfer of genetic information from the DNA to the protein. The
Genome Project has led to a huge number of available DNA sequences and,
therefore, protein sequences. The Structural Genomics initiative largely aims
to obtain “new” folds not currently present in the Protein Data Bank. Yet, the
number of available structures inevitably lags behind the number of sequences.
At the same time, an equally important problem is to find out the types and
scope of dissimilar (nonhomologous) protein sequences that adopt a similar
fold. Assembling data and comprehension of the sequence space of protein
folds should be very useful in computational protein structure prediction. This
would enhance the scope of homology modeling, which currently is the method
of choice. Thus, experimental and theoretical studies on the relationship between
sequence and structure are critical. Figuring out the relationship between sequence
and structure would further assist in the prediction of fibril structures observed in
protein misfolding diseases, and in figuring out the conformational changes
and dynamics resulting from mutations. Protein folding is one of the most
important and challenging problems in current molecular and chemical biology.
This book reviews some of the recently developed methods for studying protein
folding.
The starting point of a folding process is the unfolded state. Eliezer describes
how some of the local structural properties of the unfolded state may be charac-
terized using multidimensional nuclear magnetic resonance (NMR). Gebel and
Shortle review how the global structures of the unfolded state may be charac-
terized by measuring the residual dipolar couplings, again with NMR. Another
state that generally occurs in the process of protein folding is the rate-limiting
transition state. Pandit and coworkers describe how it can be characterized by
using the psi value analysis. Rao and coworkers employ molecular dynamics
simulation to characterize the transition state of small proteins. They propose a
technique to estimate the folding probability of those structures that are
sampled along near-equilibrium, constant temperature molecular dynamics
simulations. Bai and coworkers describe how the native-state hydrogen exchange
method could be combined with protein engineering to populate the intermediate
v
vi Preface
state for high-resolution structure determination by multidimensional NMR.
Lassalle and Akasaka describe how the high-pressure NMR technique can be
used to denature proteins and populate partially unfolded intermediates.
Thirumalai and Klimov describe the use of computer simulations and simple
models to investigate the formation of the intermediates. They show that the
equilibrium intermediates occur “on pathway” and that there is a substantial
probability that they be revisited after the native state is reached, in contrast to
kinetic intermediates. Haspel and coworkers described how a building block-
based protein folding model may be used toward a reduction of the computa-
tional complexity of protein folding. The model is based on the cutting of the
target protein sequences into building block fragments that are relatively stable
and whose conformations in solution are similar to those observed when the
fragments are chain connected in the protein molecule from which they were
derived.
One of the cutting-edge methods for studying protein folding is the single
molecule technique. Ng and coworkers show that atomic force microscopy may
be used to study the process of protein folding and unfolding. Schuler describes
how the single molecule technique coupled with fluorescence resonance transfer
may be used to study the population of proteins under equilibrium conditions
and the process of kinetic folding. The study of very fast folding process, in the
time-scale of microseconds or less, constitutes another important area in current
protein folding studies. Gai and coworkers describe the use of T-jump coupled
with infrared spectroscopy to characterize such a fast folding process. Fierz
and coworkers outline a new method that uses triplet–triplet energy transfer to
measure conformational dynamics in polypeptide chains in the unfolded
ensembles, which can set the uplimit of folding times. Streicher and Makhatadze
describe recent advances in the analysis of conformational transition in peptides
using differential scanning calorimetry. Zhou describes the replica exchange
molecular dynamics (REMD) method for enhanced sampling of the conforma-
tional space. REMD couples molecular dynamics trajectories with a temperature
exchange Monte Carlo process. Replicas are run in parallel at different tempera-
tures. This procedure allows surmounting local barriers encountered in the
simulation. REMD has proven to be a very powerful tool, and it is increasingly
used in MD simulations for small proteins/peptides. Jernigan and Kloczowski
describe a computational result that relates the packing regularities of biological
structures to their dynamics. Prediction of protein structures is the central issue of
protein folding studies. Casadio and coworkers describe how protein structures
might be predicted with and without the existence of homologs. This feat has
been achieved with the aid of machine learning-based methods specifically
suited for predicting structural features. For protein–protein interaction, a
Preface vii
knowledge-based strategy may provide predictions of putative interaction
patches on the protein surface.
It behooves us to note that not all proteins fold into stable conformations.
Recently, there has been an increasing amount of data that a large percentage
of the proteins exist in what is commonly called the natively disordered state.
This state describes a spectrum of conformations with variable stabilities. Although
on its own the stability of the native state is low (hence the term disordered),
binding to its partner enhances its stability. Thus, here too, the native state is
the functional state. Folding, misfolding, and protein disorder are all related to
each other. Progress in the understanding of one assists in comprehension of
the other. Above all, all relate to protein function and misfunction.
Altogether, Protein Folding Protocols is a comprehensive collection of
chapters describing a broad range of techniques to study, predict, and analyze
the protein folding process. It covers experiment and theory, bioinformatics
approaches, and state-of-the-art simulation protocols for better sampling of the
conformational space. Protein folding remains one of the most challenging
problems in the biological/natural sciences. Making progress in this area will
have tremendous implications, ranging from drug design, functional assignment,
comprehension of the nature of regulation, figuring out molecular machines, viral
entry into cells, and putting together cellular pathways and their dynamics. The
challenge still remains; however, experiment and theory have been making
steps toward an eventual practical solution.
Yawen Bai
Ruth Nussinov

Contents
Preface ..............................................................................................................v
Contributors .....................................................................................................xi
1. Infrared Temperature-Jump Study of the Folding Dynamics
of α-Helices and β-Hairpins
Feng Gai, Deguo Du, and Yao Xu.........................................................1
2. The Use of High-Pressure Nuclear Magnetic Resonance
to Study Protein Folding
Michael W. Lassalle and Kazuyuki Akasaka.......................................21
3. Characterization of Denatured Proteins Using Residual
Dipolar Couplings
Erika B. Gebel and David Shortle.......................................................39
4. Characterizing Residual Structure in Disordered Protein States
Using Nuclear Magnetic Resonance
David Eliezer.......................................................................................49
5. Population and Structure Determination of Hidden Folding
Intermediates by Native-State Hydrogen Exchange-Directed
Protein Engineering and Nuclear Magnetic Resonance
Yawen Bai, Hanqiao Feng, and Zheng Zhou......................................69
6. Characterizing Protein Folding Transition States Using Ψ-Analysis
Adarsh D. Pandit, Bryan A. Krantz, Robin S. Dothager,
and Tobin R. Sosnick......................................................................83
7. Advances in the Analysis of Conformational Transitions
in Peptides Using Differential Scanning Calorimetry
Werner W. Streicher and George I. Makhatadze.............................105
8. Application of Single Molecule Förster Resonance Energy
Transfer to Protein Folding
Benjamin Schuler..............................................................................115
9. Single Molecule Studies of Protein Folding
Using Atomic Force Microscopy
Sean P. Ng, Lucy G. Randles, and Jane Clarke.................................139
10. Using Triplet–Triplet Energy Transfer to Measure Conformational
Dynamics in Polypeptide Chains
Beat Fierz, Karin Joder, Florian Krieger,
and Thomas Kiefhaber..................................................................169
ix