Figure 1: Characteristic of ClyF.
(A) Time-killing curves of ClyF against S. aureus. S. aureus N315 were washed once with PBS and then treated with either 25 μg/ml ClyF, or equivalent volume of PBS buffer only, the changes of OD600nm were monitored by a microplate reader at 37 °C for 15 min (left-Y axis, blank lines). Meanwhile, the viable cell number was calculated by plating onto TSB agar plates at various times (0, 2, 5, and 15 min, right-Y axis, blue lines). The effect of temperature (B), pH (C), EDTA (left Y-axis in (D), blank line) and NaCl (right Y-axis in (D), blue line) on the lytic activity of ClyF. The lytic activity of ClyF in each condition was compared and normalized as activity relative to the maximal activity. (E–G) Storage stability of ClyF. ClyF was stored at RT (E), 4 °C (F) and −80 °C (G) for different times (0–10 months). The residual activity against S. aureus N315 was determined by a microplate reader at 37 °C for 15 min, related to the activity of the fresh ClyF before storage. (H) Circular dichroism spectra of ClyF and Pc. ClyF (7.7 μM) and Pc (10.7 μM) in 4 mM Tris-HCl (pH 7.4) were scanned by a circular dichroism spectrometer from 190–260 nm at RT. (I) Thermal denaturation curves of ClyF and Pc. ClyF (38.5 μM) and Pc (21.4 μM) in 4 mM Tris-HCl (pH 7.4) were monitored by a circular dichroism spectrometer at 220 nm at temperatures from 25–90 °C. The Tm of each protein is estimated by the instrument’s software. Groups treated with PBS were used as controls, each assay was repeated for at least three times. Mean values are plotted, and error bars represent 1× standard deviation.
