Supplementary Material

for

Compatibility of the conformationally rigid CF3Bpg side chain with the hydrophobic coiledcoil interface

Mario Salwiczek,§ Pavel Mykhailiuk,# Sergii Afonin,$ Igor Komarov,# Anne S.

Ulrich,$ Beate Koksch§,*

§Department of Biology, Chemistry, Pharmacy; Institute of Chemistry and Biochemistry; Freie Universität Berlin; Takustr. 3, 14195 Berlin, Germany

#Department of Chemistry, Kyiv National Taras Shevchenko University,

Volodymyrska 64, 01033 Kyiv, Ukraine

$Karlsruhe Institute of Technology (KIT), Institute of Biological Interfaces (IBG2) and Institute of Organic Chemistry, Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany

Phone: +49 (0)30 838 55344

Fax: +49 (0)30 838 55644

Email:

Url: http://userpage.chemie.fu-berlin.de/~akkoksch/

Folding Specificity

As shown in Figure S1-A, the isolated CF3Bpg-substituted VPK analogues adopt a random coil conformation, whereas stable a-helices are formed when mixed with equimolar amounts of the likewise random coil VPE. The equimolar mixtures were subjected to temperature induced unfolding while monitoring the CD signal at 222 nm. As shown in figure S1-B, unfolding is highly cooperative and fits well to the proposed two-state model (eq. S1).

Fig. S1 CD-spectra of the isolated CF3-Bpg-substituted VPK analogues and VPE as well as their equimolar mixtures at pH 7.4 (100 mM phosphate buffer) (A), and melting curves of the dimers (B).

For a full characterization of the original VPE-VPK coiled coil the reader is referred to Salwiczek et al. 2009.

DG = DHm · (1 – T/Tm) + DCp · [T – Tm – T · ln(T/Tm)] (S1)

As previously published (Salwiczek et al. 2009), the parallel orientation of the VPE-VPK helices has been verified applying a FRET assay. Upon mixing the N-terminally Abz-labeled (lex = 320 nm; lem = 420 nm) VPK peptides with increasing concentrations of a VPE analogue that contains an N-terminal 3-nitrotyrosine (lmax = 420 nm) as a quencher, the Abz fluorescence progressively decreases (Figure S2). The decrease in relative fluorescence intensity is comparable to the original dimer and all the other variants that have been previously investigated. The reader is referred to supporting information of Salwiczek et al. 2009.

Fig. S2 Fluorescence spectra normalized to maximum fluorescence of the CF3-Bpg-substituted VPK analogues at increasing concentrations of the quencher VPE-NYNO2 at pH 7.4 (100 mM phosphate buffer).

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