Folding and bindingThe conformational repertoire of proteins: folding,aggregation and structural recognitionEditorial overviewMikael Oliveberg and Eugene I Shakhnovich

CurrentOpinion inStructuralBiology2006,16:68–70

Available online 26th January 2006

0959-440X/$ – see front matter

Published by Elsevier Ltd.

DOI 10.1016/j.sbi.2006.01.015

Current Opinion in Structural Biology 2006, 16:68–

The protein folding field has evolved remarkably during the past several

years. Initially, following the seminal discoveries of Anfinsen, the protein

folding phenomenon was considered as belonging to the realm of biochem-

istry, whereby each protein is viewed as a unique system that requires its

own detailed characterization — akin to any specific mechanism in biology.

The introduction, in the early 1990s, of simplified models and their success

in explaining several key aspects of protein folding has shifted the thinking

of many researchers in the field towards views motivated by physics. The

‘physics’-centered approach focuses on statistical mechanical aspects of the

folding problem by emphasizing the universality of folding scenarios over

the uniqueness of folding pathways for each protein. This approach

(reviewed in [1–3]) dominated theoretical thinking during the past decade.

Its successes brought theory and experiment closer together, transforming

protein folding from a branch of biochemistry to a truly interdisciplinary

enterprise that benefits greatly from the confluence of ideas and approaches

from physics, chemistry and biology. This culminated in the resolution of

the so-called ‘Levinthal paradox’ (or, better, in the clear realization that it is

not a paradox), providing a clear physical picture of how the protein folding

problem could be solved.

Current work in the field is a quest to better understand how the protein

folding problem is actually solved in Nature. The contribution of physics-

inspired approaches to the protein folding field is twofold. First, at the level

of modeling, an improved understanding of the general principles of protein

folding makes it possible to formulate a new generation of relatively simple

yet not oversimplified models that are much more accurate than the

minimalistic lattice and off-lattice models of the past century in terms of

the structural realism of proteins and their energetics. Vastly enhanced

computer power makes it possible to carry out extensive simulations of

structurally realistic models. Also, improved understanding of the general

physical principles of protein folding allows more precise questions to be

asked and better analysis of the massive amount of data that emerge.

Second, on the experimental side, the introduction of ingenious new

approaches and enhanced interaction between theory and experiments give

valuable guidance that helps to formulate meaningful theory-inspired

hypotheses and test them experimentally. Such synergism provides an

exciting environment conducive to the rapid progress of the field.

More recently, intense attention has been focused on themolecular mechan-

isms underlying protein misfolding and aggregation processes. Studies of

protein misfolding and aggregation are important not only to better under-

Mikael Oliveberg

Department of Biochemistry and Biophysics,

Arrhenius Laboratories for Natural Sciences,

Stockholm University, 106 91 Stockholm,

Sweden

e-mail: mikael.oliveberg@dbb.su.se

Eugene I Shakhnovich

Department of Chemistry and Chemical Biology,

Harvard University, 12 Oxford Street,

Cambridge, MA 02138, USA

e-mail: eugene@belok.harvard.edu

70 www.sciencedirect.com

Editorial overview Oliveberg and Shakhnovich 69

stand the etiology and clinic of protein misfolding dis-

eases but also to make further progress towards obtaining

a fundamental understanding of the conformational prop-

erties and evolution of proteins. Of key interest is iden-

tifying the still mysterious precursor species that trigger

cytotoxic function and neural damage. Typically, but not

always, disease progression is accompanied by the deposi-

tion of macroscopic protein aggregates, indicating that the

species that cause cellular damage are oligomeric. How-

ever, it seems that it is not themature deposits that are the

primary cause of the problem, also these are morpholo-

gically different in different protein misfolding diseases,

but rather the smaller oligomers populated during the

macroscopic aggregation process [4,5]. At the level of

monomeric proteins, the noxious pathway seems to ori-

ginate from the poorly defined ensembles of disordered,

or partly disordered, states that either accumulate early in

the folding process or arise from fluctuations of the native

structure [6]. Further progress in this direction relies on

better understanding the structural properties of dena-

tured states and the conformational repertoire of native

proteins. Protein aggregation, as opposed to protein fold-

ing, is a high-order process controlled by mass transfer. As

a result, the aggregation pathway is not only controlled by

the sidechain composition, but is also highly malleable

and depends critically on the experimental conditions [7].

This structural malleability poses an extra challenge

when it comes to extrapolating in vitro data to the

crowded and geometrically confined interior of the living

cell. Finally, one of the main ‘applied’ objectives of

protein folding research is currently to provide the struc-

tural basis for the rational development of drugs to

therapeutically intervene in protein misfolding and

loss-of-function disorders. In this case, understanding

the rules of biological interactions translates into advance-

ment in drug discovery. Protein folding is thus only one

aspect of a much larger fundamental biological phenom-

enon — the specificity of structural recognition that gives

rise to specific biological shapes of molecules and their

complexes. This section features a diverse set of reviews

that reflect fully on recent progress in the field, in both

theory and experiment.

Theoretical work on protein folding dynamics is reviewed

by Caflisch. He points out that thermodynamic sampling

of the energy, entropy and potential of mean force of a

protein along pre-selected global conformational para-

meters, such as number of native contacts (Q), first

introduced and calculated in [8] and now known as the

‘folding funnel’ [9], although instructive for very idea-

lized models, may be less useful for more realistic models.

The Caflisch group and others have proposed an alter-

native approach — the protein folding network — that

does not rely on sampling along pre-determined reaction

coordinates and is capable of incorporating kinetic infor-

mation directly into conformational network analysis.

Caflisch discusses the analysis of molecular dynamics

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simulations of small b proteins and other systems using

the new approach.

A number of theoretical techniques to simulate small and

large molecules and their aggregates are reviewed by

Dokholyan. He describes such approaches as molecular

dynamics simulations and all-atom Monte Carlo techni-

ques, and focuses on a computationally efficient simula-

tionmethod, discontinuousmolecular dynamics (DMD),

which was recently implemented for protein models of

various levels of realism, including detailed all-atom

models. The author discusses how these techniques

are employed to gain insights into ‘milestones’ on the

folding pathway — intermediates and transition state

ensembles. Furthermore, simulations have recently been

applied with great success to study the aggregation and

misfolding of proteins; these studies are highlighted in

this review.

Several reviews are devoted to experimental studies of

protein folding and binding. Bilsel and Matthews discuss

experimental approaches to study the first event in pro-

tein folding. Non-specific collapse at the initial stage of

the protein folding process was uncovered by many

simulations and experiments. The authors present the

most recent results from high temporal resolution experi-

ments of initial protein collapse and discuss their signifi-

cance to understanding the general principles of protein

folding.

A new approach to study the energetics of protein folding

is reviewed by Kelly and co-authors. They point out that a

traditional protein engineering approach (based on f-

value analysis) can probe the contribution of sidechain

interactions to protein stabilization, but is less efficient for

studying the role of backbone hydrogen bonds. The

authors discuss how new methods based on the introduc-

tion of unnatural amino acids can provide detailed infor-

mation on the energetics of hydrogen bonding in b

proteins.

Swain and Gierasch review recent advances in studies of

the transitions taking place in the native basin of the

folding energy landscape: allosteric changes coupled to

ligand binding. The pathways of allosteric communica-

tion within native structures provide fundamental

knowledge about the ensemble properties of folded

proteins, as well as about the nature of the structural

transitions taking place in compact regions of conforma-

tional space. The authors discuss the role of allostery in

structural recognition and present cases in which allos-

teric transitions have been engineered into proteins to

tailor new functions.

Also shedding light on the events taking place on the

other side of the folding free energy barrier, Takeuchi and

Wagner review the development of NMR methods for

Current Opinion in Structural Biology 2006, 16:68–70

70 Folding and binding

characterizing weak protein interactions, both between

states that are largely disordered and between proteins

and small-molecule compounds. Particular emphasis is

put on binding-mediated folding processes and drug

design.

Rousseau, Schymkowitz and Serrano review recent

advances in determining the structure of protein aggre-

gates (amyloid fibrils) and the molecular determinants

controlling their formation. Aggregation-prone sequences

can now be identified by their amino acid composition

and it has become increasingly clear that natural proteins

employ anti-design to avoid erroneous association. More-

over, new results demonstrate that the pathways and

molecular determinants of different aggregation phenom-

ena differ: intervention with amyloid fibrils need not

affect alternative pathways leading to other types of

aggregates. This observation is of central importance to

obtaining a deeper understanding of the mechanism

underlying protein misfolding diseases.

A traditional paradigm in drug discovery is that small-

molecule drugs should bind to their targets as specifically

as possible. Hopkins, Mason and Overington, in

their review, offer a complementary view— an approach

based on ‘promiscuous drugs’. They show how studies

of promiscuous drugs can be of both practical importance

and fundamental value to studies of protein–small

molecule interactions and, in a broader context, studies

Current Opinion in Structural Biology 2006, 16:68–70

of the organismal effect of drugs, such as toxicity and

ADME (absorption-distribution-metabolism-excretion)

properties.

References1. Mirny L, Shakhnovich E: Protein folding theory: from lattice

to all-atom models. Annu Rev Biophys Biomol Struct 2001,30:361-396.

2. Shea JE, Brooks CL III: From folding theories to foldingproteins: a review and assessment of simulation studies ofprotein folding and unfolding. Annu Rev Phys Chem 2001,52:499-535.

3. Pande VS, Grosberg A, Tanaka T, Rokhsar DS: Pathways forprotein folding: is a new view needed? Curr Opin Struct Biol1998, 8:68-79.

4. Dobson CM: Protein folding and misfolding. Nature 2003,426:884-890.

5. Sekijima Y, Wiseman RL, Matteson J, Hammarstrom P, Miller SR,Sawkar AR, Balch WE, Kelly JW: The biological and chemicalbasis for tissue-selective amyloid disease. Cell 2005,121:73-85.

6. Thirumalai D, Klimov DK, Dima RI: Emerging ideas on themolecular basis of protein and peptide aggregation.Curr Opin Struct Biol 2003, 13:146-159.

7. Otzen DE, Oliveberg M: Transient formation of nano-crystallinestructures during fibrillation of an Ab-like peptide. Protein Sci2004, 13:1417-1421.

8. Sali A, Shakhnovich E, Karplus M: How does a protein fold?Nature 1994, 369:248-251.

9. Onuchic JN, Wolynes PG, Luthey-Schulten Z, Socci ND: Towardan outline of the topography of a realistic protein-foldingfunnel. Proc Natl Acad Sci USA 1995, 92:3626-3630.

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