Protein Folding in the Cell: Challenges and Progress This Review Comes from a Themed Issue on Folding and Binding Edited Macromolecular Crowding Hindered Mobility and Sticky Neighbors Vectorial Synthesis and Roles of Mrna and Ribosomes in Folding Concluding Thoughts

It is hard to imagine a more extreme contrast than that between the dilute solutions used for in vitro studies of protein folding and the crowded, compartmentalized, sticky, spatially inhomogeneous interior of a cell. This review highlights recent research exploring protein folding in the cell with a focus on issues that are generally not relevant to in vitro studies of protein folding, such as macromolecular crowding, hindered diffusion, cotranslational folding, molecular chaperones, and evolutionary pressures. The technical obstacles that must be overcome to characterize protein folding in the cell are driving methodological advances, and we draw attention to several examples, such as fluorescence imaging of folding in cells and genetic screens for in-cell stability.

[1]  Sen Li,et al.  Effect of Ficoll 70 on thermal stability and structure of creatine kinase , 2010, Biochemistry (Moscow).

[2]  Claus O Wilke,et al.  Signatures of protein biophysics in coding sequence evolution. , 2010, Current opinion in structural biology.

[3]  H. Bernstein,et al.  The conformation of a nascent polypeptide inside the ribosome tunnel affects protein targeting and protein folding , 2010, Molecular microbiology.

[4]  Everett A Lipman,et al.  Single-molecule spectroscopy of protein folding in a chaperonin cage , 2010, Proceedings of the National Academy of Sciences.

[5]  A. Driessen,et al.  Sec- and Tat-mediated protein secretion across the bacterial cytoplasmic membrane--distinct translocases and mechanisms. , 2008, Biochimica et biophysica acta.

[6]  Matthew I Peña,et al.  An adaptive mutation in adenylate kinase that increases organismal fitness is linked to stability-activity trade-offs. , 2007, Protein engineering, design & selection : PEDS.

[7]  Matthew I Peña,et al.  Evolutionary fates within a microbial population highlight an essential role for protein folding during natural selection , 2010, Molecular systems biology.

[8]  Ryan Hunt,et al.  Silent (synonymous) SNPs: should we care about them? , 2009, Methods in molecular biology.

[9]  M. Gerstein,et al.  Diverse Cellular Functions of the Hsp90 Molecular Chaperone Uncovered Using Systems Approaches , 2007, Cell.

[10]  Jeffrey Skolnick,et al.  Crowding and hydrodynamic interactions likely dominate in vivo macromolecular motion , 2010, Proceedings of the National Academy of Sciences.

[11]  Zoya Ignatova,et al.  Folding at the birth of the nascent chain: coordinating translation with co-translational folding. , 2011, Current opinion in structural biology.

[12]  Zoya Ignatova,et al.  Silent mutations in sight: co-variations in tRNA abundance as a key to unravel consequences of silent mutations. , 2010, Molecular bioSystems.

[13]  Lisa D. Cabrita,et al.  Protein folding on the ribosome. , 2010, Current opinion in structural biology.

[14]  Dan S. Tawfik,et al.  Stability effects of mutations and protein evolvability. , 2009, Current opinion in structural biology.

[15]  Michele Vendruscolo,et al.  Proteome-level interplay between folding and aggregation propensities of proteins. , 2010, Journal of molecular biology.

[16]  A. Horwich,et al.  Chaperonin chamber accelerates protein folding through passive action of preventing aggregation , 2008, Proceedings of the National Academy of Sciences.

[17]  A. Komar,et al.  A pause for thought along the co-translational folding pathway. , 2009, Trends in biochemical sciences.

[18]  Zoya Ignatova,et al.  Monitoring protein stability and aggregation in vivo by real-time fluorescent labeling. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[19]  Patricia L Clark,et al.  Increased incidence of rare codon clusters at 5' and 3' gene termini:implications for function , 2010, BMC Genomics.

[20]  L. Gierasch,et al.  Macromolecular crowding remodels the energy landscape of a protein by favoring a more compact unfolded state. , 2010, Journal of the American Chemical Society.

[21]  F. Hartl,et al.  Monitoring Protein Conformation along the Pathway of Chaperonin-Assisted Folding , 2008, Cell.

[22]  Dirar Homouz,et al.  Structure, function, and folding of phosphoglycerate kinase are strongly perturbed by macromolecular crowding , 2010, Proceedings of the National Academy of Sciences.

[23]  Huan‐Xiang Zhou,et al.  Macromolecular crowding and confinement: biochemical, biophysical, and potential physiological consequences. , 2008, Annual review of biophysics.

[24]  Andrey Kosolapov,et al.  Tertiary Interactions within the Ribosomal Exit Tunnel , 2009, Nature Structural &Molecular Biology.

[25]  J. Hunt,et al.  Assessment of protein folding/solubility in live cells. , 2003, Methods in molecular biology.

[26]  R. Morimoto,et al.  Biological and chemical approaches to diseases of proteostasis deficiency. , 2009, Annual review of biochemistry.

[27]  Adrian H Elcock,et al.  Models of macromolecular crowding effects and the need for quantitative comparisons with experiment. , 2010, Current opinion in structural biology.

[28]  M. Fares,et al.  The Effect of Chaperonin Buffering on Protein Evolution , 2010, Genome biology and evolution.

[29]  Patricia L Clark,et al.  Cotranslational folding increases GFP folding yield. , 2010, Biophysical journal.

[30]  M. Molinari,et al.  In and out of the ER: protein folding, quality control, degradation, and related human diseases. , 2007, Physiological reviews.

[31]  Y. Pilpel,et al.  An Evolutionarily Conserved Mechanism for Controlling the Efficiency of Protein Translation , 2010, Cell.

[32]  M. DeLisa,et al.  A rapid protein folding assay for the bacterial periplasm , 2010, Protein science : a publication of the Protein Society.

[33]  W. Skach.  Cellular mechanisms of membrane protein folding , 2009, Nature Structural &Molecular Biology.

[34]  Arthur L Horwich,et al.  Chaperonin-mediated protein folding: using a central cavity to kinetically assist polypeptide chain folding , 2009, Quarterly Reviews of Biophysics.

[35]  Matthias J. Feige,et al.  An unfolded CH1 domain controls the assembly and secretion of IgG antibodies. , 2009, Molecular cell.

[36]  Jon Beckwith,et al.  Mechanisms of oxidative protein folding in the bacterial cell envelope. , 2010, Antioxidants & redox signaling.

[37]  H. Rye,et al.  GroEL stimulates protein folding through forced unfolding , 2008, Nature Structural &Molecular Biology.

[38]  S. Cavagnero,et al.  Dynamic fluorescence depolarization: a powerful tool to explore protein folding on the ribosome. , 2010, Methods.

[39]  Zoya Ignatova,et al.  Transient ribosomal attenuation coordinates protein synthesis and co-translational folding , 2009, Nature Structural &Molecular Biology.

[40]  David Bogumil,et al.  Chaperonin-Dependent Accelerated Substitution Rates in Prokaryotes , 2010, Genome biology and evolution.

[41]  S. Radford,et al.  Optimizing protein stability in vivo. , 2009, Molecular cell.

[42]  Hidekazu Hiroaki,et al.  High-resolution multi-dimensional NMR spectroscopy of proteins in human cells , 2009, Nature.

[43]  Kurt Fredrick,et al.  How the Sequence of a Gene Can Tune Its Translation , 2010, Cell.

[44]  Margaret S. Cheung,et al.  Factors Defining Effects of Macromolecular Crowding on Protein Stability: An in Vitro/in Silico Case Study Using Cytochrome c , 2011 .

[45]  N. Dokholyan,et al.  Macromolecular crowding induces polypeptide compaction and decreases folding cooperativity. , 2010, Physical chemistry chemical physics : PCCP.

[46]  Klaus Schulten,et al.  Structural Insight into Nascent Polypeptide Chain–Mediated Translational Stalling , 2009, Science.

[47]  R. Riek,et al.  Cotranslational structure acquisition of nascent polypeptides monitored by NMR spectroscopy , 2010, Proceedings of the National Academy of Sciences.

[48]  Conggang Li,et al.  Effects of proteins on protein diffusion. , 2010, Journal of the American Chemical Society.

[49]  J. Riemer,et al.  Disulfide Formation in the ER and Mitochondria: Two Solutions to a Common Process , 2009, Science.

[50]  P. Tian,et al.  Secretion of a bacterial virulence factor is driven by the folding of a C-terminal segment , 2010, Proceedings of the National Academy of Sciences.

[51]  Sebastian Mayer,et al.  Correlation of levels of folded recombinant p53 in escherichia coli with thermodynamic stability in vitro. , 2007, Journal of molecular biology.

[52]  Eugene I Shakhnovich,et al.  Understanding protein evolution: from protein physics to Darwinian selection. , 2008, Annual review of physical chemistry.

[53]  Nora H Barakat,et al.  Exploiting elements of transcriptional machinery to enhance protein stability. , 2007, Journal of molecular biology.

[54]  J. Beckwith,et al.  Detecting Folding Intermediates of a Protein as It Passes through the Bacterial Translocation Channel , 2009, Cell.

[55]  David Tollervey,et al.  Coding-Sequence Determinants of Gene Expression in Escherichia coli , 2009, Science.

[56]  Yutaka Ito,et al.  Cellular structural biology. , 2010, Current opinion in structural biology.

[57]  Conggang Li,et al.  Translational and rotational diffusion of a small globular protein under crowded conditions. , 2009, The journal of physical chemistry. B.

[58]  Colin J. Daniel,et al.  Control of Translocation through the Sec61 Translocon by Nascent Polypeptide Structure within the Ribosome*♦ , 2008, Journal of Biological Chemistry.

[59]  J. Kappes,et al.  A Synonymous Single Nucleotide Polymorphism in ΔF508 CFTR Alters the Secondary Structure of the mRNA and the Expression of the Mutant Protein* , 2010, The Journal of Biological Chemistry.

[60]  F. Hartl,et al.  Chaperonin-Catalyzed Rescue of Kinetically Trapped States in Protein Folding , 2010, Cell.

[61]  H. Taguchi,et al.  A systematic survey of in vivo obligate chaperonin‐dependent substrates , 2010, The EMBO journal.

[62]  Vectorial Transport and Folding of an Autotransporter Virulence Protein During Outer Membrane Secretion , 2009 .

[63]  U. Mortensen,et al.  A versatile selection system for folding competent proteins using genetic complementation in a eukaryotic host , 2010, Protein science : a publication of the Protein Society.

[64]  Andrew C. Miklos,et al.  Protein nuclear magnetic resonance under physiological conditions. , 2009, Biochemistry.

[65]  D. Hebert,et al.  The Cotranslational Maturation Program for the Type II Membrane Glycoprotein Influenza Neuraminidase* , 2008, Journal of Biological Chemistry.

[66]  C. Deutsch,et al.  A folding zone in the ribosomal exit tunnel for Kv1.3 helix formation. , 2010, Journal of molecular biology.

[67]  Bernd Bukau,et al.  The ribosome as a platform for co-translational processing, folding and targeting of newly synthesized proteins , 2009, Nature Structural &Molecular Biology.

[68]  H. Saibil,et al.  Chaperonin complex with a newly folded protein encapsulated in the folding chamber , 2009, Nature.

[69]  Huan‐Xiang Zhou,et al.  Atomistic modeling of macromolecular crowding predicts modest increases in protein folding and binding stability. , 2009, Biophysical journal.

[70]  Daniel N. Wilson,et al.  Structural basis for translational stalling by human cytomegalovirus and fungal arginine attenuator peptide. , 2010, Molecular cell.

[71]  Zoya Ignatova,et al.  From the test tube to the cell: exploring the folding and aggregation of a beta-clam protein. , 2007, Biopolymers.

[72]  M. Vendruscolo,et al.  Towards quantitative predictions in cell biology using chemical properties of proteins. , 2008, Molecular bioSystems.

[73]  Johannes Buchner,et al.  How antibodies fold. , 2010, Trends in biochemical sciences.

[74]  K. Inaba.  Disulfide bond formation system in Escherichia coli. , 2009, Journal of biochemistry.

[75]  Nora H Barakat,et al.  Combined use of experimental and computational screens to characterize protein stability. , 2010, Protein engineering, design & selection : PEDS.

[76]  J. D. Mcdonald,et al.  Protein folding stability and dynamics imaged in a living cell , 2010, Nature Methods.

[77]  T. Terwilliger,et al.  New Molecular Reporters for Rapid Protein Folding Assays , 2008, PloS one.

[78]  Anja Nenninger,et al.  Size Dependence of Protein Diffusion in the Cytoplasm of Escherichia coli , 2010, Journal of bacteriology.

[79]  Titus M. Franzmann,et al.  Protein refolding by pH-triggered chaperone binding and release , 2009, Proceedings of the National Academy of Sciences.

[80]  S. Ghaemmaghami,et al.  Quantitative protein stability measurement in vivo , 2001, Nature Structural Biology.

[81]  B. Freeman,et al.  Slowing bacterial translation speed enhances eukaryotic protein folding efficiency. , 2010, Journal of molecular biology.

[82]  Koreaki Ito,et al.  The Ribosomal Exit Tunnel Functions as a Discriminating Gate , 2002, Cell.

[83]  S. Hagen,et al.  Solvent viscosity and friction in protein folding dynamics. , 2010, Current protein & peptide science.

[84]  D. Sideris,et al.  Oxidative protein folding in the mitochondrial intermembrane space. , 2010, Antioxidants & redox signaling.

[85]  Mohit Kumar,et al.  Mobility of cytoplasmic, membrane, and DNA-binding proteins in Escherichia coli. , 2010, Biophysical journal.

[86]  Eytan Ruppin,et al.  Translation efficiency is determined by both codon bias and folding energy , 2010, Proceedings of the National Academy of Sciences.

[87]  Adrian H. Elcock,et al.  Diffusion, Crowding & Protein Stability in a Dynamic Molecular Model of the Bacterial Cytoplasm , 2010, PLoS Comput. Biol..

[88]  R. Glockshuber,et al.  FRET-based in vivo screening for protein folding and increased protein stability. , 2003, Journal of molecular biology.

[89]  E. Orlova,et al.  Topologies of a Substrate Protein Bound to the Chaperonin GroEL , 2007, Molecular cell.