Melting temperature

Stability/Folding

Melting Temperature (Tm)

°C (Celsius)

Circular Dichroism (CD)

5 mM PBS

25-100℃

7.0

0.1 mg/ml

215 nm

None

The secondary structure and protein stability analysis by circular dichroism (CD) was performed using a Jasco J-715 spectropolarimeter (Jasco Inc., Easton, MD). The purified proteins were dialyzed against 5 mM phosphate buffered saline (PBS) (68.5 mM NaCl, 1 mM KCl, 5 mM phosphate, pH 7.0) and diluted to a concentration of 0.1 mg/ml. For thermal denaturation analysis, protein samples were heated from 25 °C to 100 °C in 1 °C increments while monitoring CD ellipticity at 215 nm. To facilitate comparison, raw data were converted to fraction unfolded and analyzed using a two-state transition model (5, 6). The data shown in Fig. S3 were fitted to sigmoidal curves using Prism 6 (GraphPad Software, Inc., LaJolla, CA), yielding the melting temperatures Tm reported in Table 1 of the main text.

References:
5. Minor DL, Jr., Kim PS. 1994. Measurement of the β-sheet-forming propensities of amino acids.
Nature 367:660-663. http://dx.doi.org/10.1038/367660a0.
6. Smith CK, Withka JM, Regan L. 1994. A thermodynamic scale for the β-sheet forming tendencies
of the amino acids. Biochemistry 33:5510-5517.