α-Helix Peptide Folding and Unfolding Activation Barriers:  A Nanosecond UV Resonance Raman Study

Lednev, Igor K.; Karnoup, Anton S.; Sparrow, Mark C.; Asher, Sanford A.

doi:10.1021/ja991382f

Cited by 207 publications

(539 citation statements)

References 90 publications

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“…46,47 Therefore, the two-state assumption has been used in T-jump studies in the past. 7,48 Herein, to compare the folding kinetics of any two peptides, we may also invoke the two-state model below to describe their relaxation kinetics.…”

Section: Discussionmentioning

confidence: 99%

“…15,16,18,21,22,[28][29][30][31] Experimentally, rapid laser-induced T-jump methods have allowed the first investigations of the formation of -helices. [4][5][6][7][8][9][10][11] In these studies, a rapid T-jump pulse induces a shift in the equilibrium between the coil and helix states, and the subsequent relaxation toward the new thermal equilibrium point is probed by examining all of the amide groups via either IR 4,8,11 or Raman 7 spectroscopy, or individual residues by fluorescence spectroscopy 5 or isotopeediting 9,10 techniques. However, the peptides used in these studies are rather similar.…”

Section: Introductionmentioning

confidence: 99%

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Length Dependent Helix−Coil Transition Kinetics of Nine Alanine-Based Peptides

et al. 2004

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It is well-known that end caps and the peptide length can dramatically influence the thermodynamics of the helix-coil transition. However, their roles in determining the kinetics of the helix-coil transition have not been studied extensively and are less well understood. Kinetic Ising models and sequential kinetic models involving barrier crossing via diffusion all predict that the helix formation time depends monotonically on the peptide length with the relaxation time increasing with respect to increasing chain length. Here, we have studied the helix-coil transition kinetics of a series of Ala-based -helical peptides of different length (19-39 residues), with and without end caps, using time-resolved infrared spectroscopy coupled with laser-induced temperature jump (T-jump) initiation method. The helical content of these peptides was kinetically monitored by probing the amide carbonyl stretching frequencies (i.e., the amide I band) of the peptide backbone. We found that the relaxation rates for peptides with efficient end caps are more rapid than those of the corresponding peptides without good end caps. These results indicate that efficient end-capping sequences can not only stabilize preexisting helices but also promote helix formation through initiation. Furthermore, we found that the relaxation times of these peptides, following a T-jump of 1-11 °C, show rather complex behaviors as a function of the peptide length, in disagreement with theoretical predications. Theses results are not readily explained by theories in which Ala is taken to have a single helical propensity (s). However, recent studies have suggested that s depends on chain length; when this factor is considered, the mean first-passage times of the coil-to-helix transition show similar dependence on the peptide length as those observed experimentally.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Length Dependent Helix−Coil Transition Kinetics of Nine Alanine-Based Peptides

et al. 2004

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“…Typical R-helical peptides show an AmIII 3 band maximum at 1263 cm -1 . The 9 cm -1 shift in the AmIII 3 band position of gp41 659-671 as compared to typical R-helical peptides such as the AP peptide (24) indicates the average ψ angle of gp41 [659][660][661][662][663][664][665][666][667][668][669][670][671] deviates from the typical R-helix conformation by 10°. This result demonstrates that UVRR can discriminate between conformationally similar R-and 3 10 -helicies.…”

Section: Conformation Analysismentioning

confidence: 99%

UV Resonance Raman Investigation of a 3₁₀-Helical Peptide Reveals a Rough Energy Landscape

Ahmed

Asher

2006

Biochemistry

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We used UVRRS at 194 and 204 nm excitation to examine the backbone conformation of a 13-residue polypeptide (gp41 [659][660][661][662][663][664][665][666][667][668][669][670][671] ) that has been shown by NMR to predominantly fold into a 3 10 -helix. Examination of the conformation sensitive AmIII 3 region indicates the peptide has significant populations of -turn, PPII, 3 10 -helix, and π-helix-like conformations but little R-helix. We estimate that at 1°C on average six of the 12 peptide bonds are in folded conformations (predominantly 3 10 -and π-helix), while the other six are in unfolded ( -turn/PPII) conformations. The folded and unfolded populations do not change significantly as the temperature is increased from 1 to 60°C, suggesting a unique energy landscape where the folded and unfolded conformations are essentially degenerate in energy and exhibit identical temperature dependences.An understanding of the mechanisms of protein folding will enable the de novo design of proteins with profound commercial and medical applications (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19). Over the past 50 years, significant effort to elucidate protein folding and unfolding mechanisms has been expended. Part of this effort has examined the thermodynamics and kinetics of small model systems such as -hairpins (20)(21)(22) and alanine-based R-helices (23-33). These studies have provided detailed microscopic knowledge of the folding energy landscape of these isolated motifs. Theoretical simulations have suggested that 3 10 -helices and/or type III -turns may serve as intermediates in the R-helix folding pathway (34-39). The formation of kinetic intermediates such as -turn or 3 10 -helix lowers the entropic cost of nucleating the first helical residue, thus facilitating the helix folding transition (34,40,41).The 3 10 -helix is the fourth most common secondary structural motif in proteins (34-59). On average, in proteins, 3-4% of peptide residues occur in a 3 10 -helix conformation (48). The 3 10 -helix differs from an R-helix in that the tighter packing of the backbone forces the CdO‚‚‚H-N hydrogen bonds to point outward, away from the helical axis, resulting in decreased stability of the 3 10 -helix compared to that of the R-helix (25,48).Recent ESR (39, 42) and NMR (43) work by Millhauser et al. (41) suggests the presence of 3 10 -helix at the terminus of short alanine-based helical peptides. The authors proposed that 3 10 -helices are a relic of the folding process. They reasoned that nascent helices contain mixtures of unfolded and turn conformations, specifically type III turns. As folding conditions begin to favor helical structures, the type III turn conformation (a single 3 10 -helix unit) propagates; i.e., the chain adopts a 3 10 -helix conformation. As the helical domain lengthens, the R-helix conformation competes with the 3 10 -helix conformation. Finally, at longer helical lengths, the R-helix conformation dominates. Some 3 10 -helix/type III turn-like conformations may survive...

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“…To date, synthetic schemes have been successfully developed for the synthesis of R-helix-forming peptides, and much has been uncovered about their folding dynamics (6)(7)(8)(9)(10)(11)(12)(13). In contrast, only limited progress has been made toward understanding the synthesis of and the dynamics of -sheet peptides.…”

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confidence: 99%

UV Raman Studies of Peptide Conformation Demonstrate That Betanova Does Not Cooperatively Unfold

Boyden

Asher

2001

Biochemistry

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We used UV resonance Raman spectroscopy (UVRR) excited within the peptide bond π f π* electronic transitions and within the aromatic amino acid π f π* electronic transitions to examine the temperature dependence of the solution conformation of betanova, a 20-residue -sheet polypeptide [Kortemme, T., Ramirez-Alvarado, M., and Serrano, L. (1998) Science 281, 253-256]. The 206.5 nm excited UVRR enhances the amide vibrations and demonstrates that betanova has a predominantly -sheet structure between 5 and 82°C. The 229 nm excited UVRR, which probes the tyrosine and tryptophan side chain vibrations, shows an increase in the solvent exposure of the tryptophan side chains as the temperature is increased. Our results are consistent with the existence of an intermediate state similar to that calculated by Bursulaya and Brooks [Bursulaya, B. D., and Brooks, C. L. (1999) J. Am. Chem. Soc. 121, 9947-9951] and exclude the previously proposed two-state cooperative folding mechanism. Betanova's structure appears to be molten globule over the 3-82°C temperature range of our study.The completion of the human genome sequence makes available the primary sequence of all proteins synthesized by the human body. This represents only the first stage in the development of an understanding of the protein structural basis of disease. The next stage in this development requires an ability to predict the structure and function of proteins from their primary sequences. At present this is impossible, since not enough is known about the dynamics and thermodynamics of protein folding (1-5).The understanding of protein folding mechanisms is advancing as experimentalists develop better strategies for the de novo design of model peptides and better methodologies for characterizing equilibrium secondary structures, as well as the dynamics of folding. To date, synthetic schemes have been successfully developed for the synthesis of R-helix-forming peptides, and much has been uncovered about their folding dynamics (6-13). In contrast, only limited progress has been made toward understanding the synthesis of and the dynamics of -sheet peptides. Primarily, studies have focused on -hairpins rather than complete sheet structures (14-18). Successful synthesis of three-stranded -sheet structures has been accomplished only recently. However, these model peptides only form complete -sheet structures under limited conditions, in the presence of organic solvents and with the use of unnatural amino acids (19)(20)(21)(22). Thus, there remains much to learn about -sheet folding.Significant advances have also occurred in the theoretical understanding of protein folding (23-28). Such work includes molecular dynamics simulations of energy landscapes that describe folding thermodynamics. Currently, some of these models accurately predict experimental results. Thus, theory can now impact how experimental data are interpreted. It seems likely that the most powerful insights into the rules of protein folding will result from combined experimental and theoreti...

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α-Helix Peptide Folding and Unfolding Activation Barriers: A Nanosecond UV Resonance Raman Study

Cited by 207 publications

References 90 publications

Length Dependent Helix−Coil Transition Kinetics of Nine Alanine-Based Peptides

Length Dependent Helix−Coil Transition Kinetics of Nine Alanine-Based Peptides

UV Resonance Raman Investigation of a 3₁₀-Helical Peptide Reveals a Rough Energy Landscape

UV Raman Studies of Peptide Conformation Demonstrate That Betanova Does Not Cooperatively Unfold

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α-Helix Peptide Folding and Unfolding Activation Barriers: A Nanosecond UV Resonance Raman Study

Cited by 207 publications

References 90 publications

Length Dependent Helix−Coil Transition Kinetics of Nine Alanine-Based Peptides

Length Dependent Helix−Coil Transition Kinetics of Nine Alanine-Based Peptides

UV Resonance Raman Investigation of a 310-Helical Peptide Reveals a Rough Energy Landscape

UV Raman Studies of Peptide Conformation Demonstrate That Betanova Does Not Cooperatively Unfold

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UV Resonance Raman Investigation of a 3₁₀-Helical Peptide Reveals a Rough Energy Landscape