Time-resolved temperature-jump infrared spectroscopy of peptides with well-defined secondary structure: A Trpzip β-hairpin variant as an example

Cytochrome c unfolds locally and reversibly upon heating at pH 3. UV resonance Raman (UVRR) spectra reveal that instead of producing unordered structure, unfolding converts turns and some helical elements to β-sheet. It also disrupts the Met80-heme bond, and was earlier shown to induce peroxidase activity. Aromatic residues that are H-bonded to a heme propionate (Trp59 and Tyr48) alter their orientation, indicating heme displacement. T-jump/UVRR measurements give time constants of 0.2, 3.9 and 67 µs for successive phases of β-sheet formation and concomitant reorientation of Trp59. UVRR spectra reveal protonation of histidines, and specifically of His26, whose H-bond to Pro44 anchors the 40s Ω loop; this loop is known to be the least stable ‘foldon’ in the protein. His26 protonation is proposed to disrupt its H-bond with Pro44, triggering the extension of a short β-sheet segment at the ‘neck’ of the 40s Ω loop into the loop itself and back into the 60’s and 70’s helices. The secondary structure change displaces the heme via H-bonds from residues in the growing β-sheet, thereby exposing it to exogenous ligands, and inducing peroxidase activity. This unfolding mechanism may play a role in cardiolipin peroxidation by cyt c during apoptosis.

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“…T-jump/UVRR spectra show this process to occur in microseconds, consistent with unfolding rates in other small proteins. 70–77 …”

Section: Discussionmentioning

confidence: 99%

His26 Protonation in Cytochrome c Triggers Microsecond β-Sheet Formation and Heme Exposure: Implications for Apoptosis

Balakrishnan

Spiro

2012

J. Am. Chem. Soc.

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“…40 This analysis leads to an estimated T m ≈ 72.6°C for the denaturation temperature in plain buffer, in agreement with values of T m ≈ 72-75°C typically reported in CD studies of TZ2. [26][27][28][29] In order to enhance the Tjump relaxation signal near room temperature, we suppressed the folding midpoint slightly by adding a moderate amount of denaturant (2 M GdnHCl). This does not greatly affect the heterogeneous folding properties of TZ2.…”

Section: Methodsmentioning

confidence: 99%

“…The TZ2 hairpin is notable for its stability: Circular dichroism (CD) measurements indicate a melting temperature T m ≈ 72-75°C. [26][27][28][29][30] However, its folding is heterogeneous in the sense that different spectroscopic probes, including fluorescence emission intensity, peak fluorescence wavelength, and amide-I′ spectroscopy, report different melting temperatures in equilibrium unfolding studies. 28,30,31 For example, while CD indicates unfolding near 70°C, twodimensional infrared (IR) studies show that the native hydrogen bonds between the two strands persist up to very high temperatures of N 80°C.…”

Section: Introductionmentioning

confidence: 99%

Solvent Friction Changes the Folding Pathway of the Tryptophan Zipper TZ2

Narayanan

Pelakh

Hagen

2009

Journal of Molecular Biology

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“…T-jump experiments were performed on the home-built instrument which has been described in detail previously [29]. The temperature jump in the sample was obtained by a Raman-shifted 5 ns Nd: YAG laser pulse at 1909 nm.…”

Section: T-jump Measurementsmentioning

confidence: 99%

Impact of β‐Turn Sequence on β‐Hairpin Dynamics Studied with Infrared‐Detected Temperature Jump

Popp

Keiderling

et al. 2012

Journal of Spectroscopy

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Abstract. Folding dynamics for β-structure loss and disordered structure gain were studied in a model β-hairpin peptide based on Cochran's tryptophan zipper peptide Trpzip2, but with an altered Thr-Gly (TG) turn sequence, that is, SWTWETGKWTWK, using laser-induced temperature-jump (T-jump) kinetics with IR detection. As has been shown previously, the TG turn sequence reduces the thermodynamic β-hairpin stability as compared to the Asn-Gly sequence used in Trpzip2 (TZ2-NG). In this study, we found that the TG-turn slows down the overall relaxation dynamics as compared to TZ2-NG, which were studied at higher temperatures where the time constants show little difference between relaxation of the β-strand and the disordered conformation. These time constants become equivalent at lower temperatures for TZ2-TG than was seen for TZ2-NG. The correlation of thermodynamic stability and rates of relaxation suggests that the change from NG to TG turn results in a slowing of folding, lower k f , with less change of the unfolding rate, ku, assuming two state behavior at higher temperatures.

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Time-resolved temperature-jump infrared spectroscopy of peptides with well-defined secondary structure: A Trpzip β-hairpin variant as an example

Cited by 30 publications

References 43 publications

His26 Protonation in Cytochrome c Triggers Microsecond β-Sheet Formation and Heme Exposure: Implications for Apoptosis

His26 Protonation in Cytochrome c Triggers Microsecond β-Sheet Formation and Heme Exposure: Implications for Apoptosis

Solvent Friction Changes the Folding Pathway of the Tryptophan Zipper TZ2

Impact of β‐Turn Sequence on β‐Hairpin Dynamics Studied with Infrared‐Detected Temperature Jump

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