Figure 1. Quality control of the purified non-myristoylated CHP3 and myristoylated CHP3 (mCHP3).
(A) Sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) analysis (16%, tricine mini gels) showed a mobility shift of mCHP3 in comparison to CHP3; positions of co-migrated molecular mass standards are indicated in kDa on the left; Figure 1—source data 1: Full gel for (A). (B) Left, deconvoluted mass spectra of CHP3 (top) and mCHP3 (bottom) show a measured intact protein mass of 24.617 and 24.828 kDa (∆Mr = 211 Da), which is in line with the addition of the myristoyl group of 210 Da. For mCHP3, also low abundant dimeric species were observed by native MS analysis. Right, zoom-in spectra in the range of 24.8 ± 0.3 kDa that show differences in masses as indicated by dashed lines. In the presence of Ca2+, the measured intact protein mass of CHP3 and mCHP3 was increased by ~40 Da (mass accuracy ± 1 Da), respectively, indicating binding of one Ca2+ ion. (C) Ion mobility arrival time distributions for the +8 charge state of CHP3 (solid line) and mCHP3 (dashed line). A shoulder with shorter arrival times indicates the presence of a low abundant dimeric species for CHP3 and mCHP3. Arrival times of mCHP3 were reduced by 1.36 ms (±0.37 ms; n = 3; p = 0.024) compared to CHP3. See Figure 1—figure supplement 2 for non-deconvoluted mass spectra showing charge state distributions of CHP3 and mCHP3 measured in the positive ion mode by native MS.
Figure 1—figure supplement 1. Analytical size exclusion chromatography of purified CHP3 (A) and mCHP3 (B) on Superdex 75 Increase 10/300 GL.
Figure 1—figure supplement 2. Non-deconvoluted mass spectra showing charge state distributions of CHP3 and mCHP3 measured in the positive ion mode by native MS.
