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. 2012 Jan 26;116(3):1134-42.
doi: 10.1021/jp208918n. Epub 2012 Jan 10.

UV Resonance Raman Studies of the NaClO4 Dependence of poly-L-lysine Conformation and Hydrogen Exchange Kinetics

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UV Resonance Raman Studies of the NaClO4 Dependence of poly-L-lysine Conformation and Hydrogen Exchange Kinetics

Lu Ma et al. J Phys Chem B. .
Free PMC article

Abstract

We used 204 nm excitation UV Resonance Raman (UVRR) spectroscopy to examine the effects of NaClO(4) on the conformation of poly-L-lysine (PLL). The presence of NaClO(4) induces the formation of α-helix, π-helix/bulge, and turn conformations. The dependence of the AmIII(3) frequency on the peptide Ψ Ramachandran angle allows us to experimentally determine the conformational population distributions and the energy landscape of PLL along the Ramachandran Ψ angle. We also used UVRR to measure the NaClO(4) concentration dependence of PLL amide hydrogen exchange kinetics. Exchange rates were determined by fitting the D(2)O exchanging PLL UVRR AmII' band time evolution. Hydrogen exchange is slowed at high NaClO(4) concentrations. The PLL AmII' band exchange kinetics at 0.0, 0.2, and 0.35 M NaClO(4) can be fit by single exponentials, but the AmII' band kinetics of PLL at 0.8 M NaClO(4) requires a double exponential fit. The exchange rates for the extended conformations were monitored by measuring the C(α)-H band kinetics. These kinetics are identical to those of the AmII' band until 0.8 M NaClO(4) whereupon the extended conformation exchange becomes clearly faster than that of the α-helix-like conformations. Our results indicate that ClO(4)(-) binds to the PLL backbone to protect it from OH(-) exchange catalysis. In addition, ClO(4)(-) binding also slows the conformational exchange between the extended and α-helix-like conformations, probably by increasing the activation barriers for conformational interchanges.

Figures

Figure 1
Figure 1
(A) 204 nm UVRR spectra of PLL at pH 3 and 20 °C at 0, 0.1, 0.2, 0.35, 0.5 and 0.8 M NaClO4 concentrations. (B) Deconvolution of the 20 °C 204 nm UVRR PLL spectrum in the absence of NaClO4 at pH 3.
Figure 2
Figure 2
NaClO4 concentration dependence of 0.87 mg/ml PLL CD spectra at pH 3 and 20 °C.
Figure 3
Figure 3
Comparison of PLL α–helix–like conformation fraction calculated from CD and UVRR at different NaClO4 concentrations at pH 3 and 20 °C.
Figure 4
Figure 4
NaClO4 concentration dependence of the AmIII region of the 204 nm UVRR spectra of the calculated α–helix–like PLL conformations at pH 3 and 20 °C. See text for details.
Figure 5
Figure 5
NaClO4 concentration dependence of the Ramachandran Ψ angle distribution of PLL at pH 3 and 20 °C, assuming the T2 conformation at Ψ = 114°. The possible Ψ = 14° conformation is not shown. The extended structure Ψ angles are assumed to be those of PLL in the absence of NaClO4 as in Fig. 1B.
Figure 6
Figure 6
Clculated Gibbs free energy landscape for PLL at different NaClO4 concentrations at pH 3 and 20 °C. The dotted lines show the turn regions, assuming that the T2 turn occurs at Ψ =114°.
Figure 7
Figure 7
204 nm UVRR spectra of PLL in the absence and presence of 0.8 M NaClO4 in H2O and D2O at pH 3/pD 3 and at 20 °C.
Figure 8
Figure 8
(A) pH 2.8 UVRR of PLL at 20 °C hydrogen exchange at 28, 33, 43, and 103 s after H2O addition in the absence of NaClO4. (B) pH 2.8 UVRR PLL hydrogen exchange in 0.8 M NaClO4 obtained at 3, 8, 12, 22, 32, 42, 64, 102 and 150 min after 0.8 M NaClO4 H2O solution addition.
Figure 9
Figure 9
Time dependence of normalized AmII′ band intensity of PLL due to H–exchange at pH 2.8 at 20 °C at different NaClO4 concentrations. The thin solid lines show single exponential fitting.
Figure 10
Figure 10
Comparison of IN,C-H (t) (blue) and 1–IN,AmII (t) (red), exchange kinetics of the Cα–H b band and AmII′ band at pH 2.8 at 20 °C. The black lines are single exponential (0.2, 0.35 M) and double exponential (0.8 M) fitting curves.
Figure 11
Figure 11
NaClO4 concentration dependence of extended PLL exchange rates. The solid line shows the eqn. 6 fitting result.
Figure 12
Figure 12
Interactions between ClO4 and PLL in the PPII conformation to protect the peptide NH from exchange. The ClO4 ion binds to the amide NH and neighboring lysine −NH3+ group to form a ring structure.

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