Influence of denatured and intermediate states of folding on protein aggregation

We simulate the aggregation thermodynamics and kinetics of proteins L and G, each of which self‐assembles to the same β/β topology through distinct folding mechanisms. We find that the aggregation kinetics of both proteins at an experimentally relevant concentration exhibit both fast and slow aggregation pathways, although a greater proportion of protein G aggregation events are slow relative to those of found for protein L. These kinetic differences are correlated with the amount and distribution of intrachain contacts formed in the denatured state ensemble (DSE), or an intermediate state ensemble (ISE) if it exists, as well as the folding timescales of the two proteins. Protein G aggregates more slowly than protein L due to its rapidly formed folding intermediate, which exhibits native intrachain contacts spread across the protein, suggesting that certain early folding intermediates may be selected for by evolution due to their protective role against unwanted aggregation. Protein L shows only localized native structure in the DSE with timescales of folding that are commensurate with the aggregation timescale, leaving it vulnerable to domain swapping or nonnative interactions with other chains that increase the aggregation rate. Folding experiments that characterize the structural signatures of the DSE, ISE, or the transition state ensemble (TSE) under nonaggregating conditions should be able to predict regions where interchain contacts will be made in the aggregate, and to predict slower aggregation rates for proteins with contacts that are dispersed across the fold. Since proteins L and G can both form amyloid fibrils, this work also provides mechanistic and structural insight into the formation of prefibrillar species.

[1]  D. Baker,et al.  Contact order, transition state placement and the refolding rates of single domain proteins. , 1998, Journal of molecular biology.

[2]  C. Dobson Protein Folding and Disease: a view from the first Horizon Symposium , 2003, Nature Reviews Drug Discovery.

[3]  Teresa Head-Gordon,et al.  Matching simulation and experiment (extended abstract): a new simplified model for simulating protein folding , 2000, RECOMB '00.

[4]  David Baker,et al.  Simple physical models connect theory and experiment in protein folding kinetics. , 2002, Journal of molecular biology.

[5]  T. Head-Gordon,et al.  Minimalist models for protein folding and design. , 2003, Current opinion in structural biology.

[6]  L. Regan,et al.  A systematic exploration of the influence of the protein stability on amyloid fibril formation in vitro. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[7]  D Baker,et al.  The sequences of small proteins are not extensively optimized for rapid folding by natural selection. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[8]  M. Hecht,et al.  Nature disfavors sequences of alternating polar and non-polar amino acids: implications for amyloidogenesis. , 2000, Journal of molecular biology.

[9]  A. Fink Protein aggregation: folding aggregates, inclusion bodies and amyloid. , 1998, Folding & design.

[10]  A. Fersht Optimization of rates of protein folding: the nucleation-condensation mechanism and its implications. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[11]  J. King,et al.  Thermolabile folding intermediates: inclusion body precursors and chaperonin substrates , 1996, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[12]  L Serrano,et al.  Similarities between the spectrin SH3 domain denatured state and its folding transition state. , 2000, Journal of molecular biology.

[13]  C M Dobson,et al.  Designing conditions for in vitro formation of amyloid protofilaments and fibrils. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[14]  K. Lindorff-Larsen,et al.  Parallel protein-unfolding pathways revealed and mapped , 2003, Nature Structural Biology.

[15]  D Baker,et al.  Kinetics of folding of the IgG binding domain of peptostreptococcal protein L. , 1997, Biochemistry.

[16]  P. Roller,et al.  Scrapie amyloid (prion) protein has the conformational characteristics of an aggregated molten globule folding intermediate. , 1994, Biochemistry.

[17]  J. King,et al.  Frequencies of amino acid strings in globular protein sequences indicate suppression of blocks of consecutive hydrophobic residues , 2001, Protein science : a publication of the Protein Society.

[18]  D. Baker,et al.  NMR characterization of residual structure in the denatured state of protein L. , 2000, Journal of molecular biology.

[19]  Soon-Ho Park,et al.  Folding dynamics of the B1 domain of protein G explored by ultrarapid mixing , 1999, Nature Structural Biology.

[20]  Soon-Ho Park,et al.  An early intermediate in the folding reaction of the B1 domain of protein G contains a native-like core. , 1997 .

[21]  C. Dobson Protein folding and misfolding , 2003, Nature.

[22]  D. Baker,et al.  Critical role of β-hairpin formation in protein G folding , 2000, Nature Structural Biology.

[23]  A. Fersht,et al.  Formation of short-lived protein aggregates directly from the coil in two-state folding. , 1999, Biochemistry.

[24]  Sheena E. Radford,et al.  Im7 folding mechanism: misfolding on a path to the native state , 2002, Nature Structural Biology.

[25]  T. Head-Gordon,et al.  Intermediates and the folding of proteins L and G , 2004, Protein science : a publication of the Protein Society.

[26]  L. Kay,et al.  NOE data demonstrating a compact unfolded state for an SH3 domain under non-denaturing conditions. , 1999, Journal of molecular biology.

[27]  V. Pande,et al.  On the transition coordinate for protein folding , 1998 .

[28]  M. Hecht,et al.  De novo amyloid proteins from designed combinatorial libraries. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[29]  E D Clark,et al.  Protein refolding for industrial processes. , 2001, Current opinion in biotechnology.

[30]  Jeanette Tångrot,et al.  Complete change of the protein folding transition state upon circular permutation , 2002, Nature Structural Biology.

[31]  C. Dobson,et al.  Stabilisation of alpha-helices by site-directed mutagenesis reveals the importance of secondary structure in the transition state for acylphosphatase folding. , 2000, Journal of molecular biology.

[32]  D. Ferguson,et al.  Simulated Annealing—Optimal Histogram Methods , 1999 .

[33]  A. Horwich Protein aggregation in disease: a role for folding intermediates forming specific multimeric interactions. , 2002, The Journal of clinical investigation.

[34]  S Doniach,et al.  Association-induced folding of globular proteins. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[35]  David Baker,et al.  A phage display system for studying the sequence determinants of protein folding , 1995, Protein science : a publication of the Protein Society.

[36]  C. Dobson,et al.  Protein aggregation and amyloid fibril formation by an SH3 domain probed by limited proteolysis. , 2003, Journal of molecular biology.

[37]  H. Roder,et al.  An early intermediate in the folding reaction of the B1 domain of protein G contains a native-like core. , 1997, Biochemistry.

[38]  R. Swendsen,et al.  THE weighted histogram analysis method for free‐energy calculations on biomolecules. I. The method , 1992 .

[39]  T. Kiefhaber,et al.  Hammond behavior versus ground state effects in protein folding: evidence for narrow free energy barriers and residual structure in unfolded states. , 2003, Journal of molecular biology.

[40]  KumarShankar,et al.  The weighted histogram analysis method for free-energy calculations on biomolecules. I , 1992 .

[41]  Sheena E. Radford,et al.  Ultrarapid mixing experiments reveal that Im7 folds via an on-pathway intermediate , 2001, Nature Structural Biology.

[42]  D Baker,et al.  Contrasting roles for symmetrically disposed beta-turns in the folding of a small protein. , 1997, Journal of molecular biology.