Fig 8. Validation of the CwlD:GerS salt bridge by co-immunoprecipitation.
(A) Western blot of GerS in co-immunoprecipitation fractions obtained from pulling down FLAG-tagged proteins produced in sporulating cells for wild-type CwlD, CwlDR169D, CwlDH49A, or CwlDE199A. CwlD was detected using an anti-FLAG antibody, while untagged GerS was detected using an anti-GerS antibody. CwlD-c and GerS-c indicate cleaved form of both proteins. (B) Percentage of GerS pulled down with FLAG-tagged CwlD relative to the amount detected in pull-downs with FLAG-tagged wild-type CwlD. A mean of three independent assays is shown, except for CwlDE199A-FLAG, for which two independent biological replicates were performed. Dots represent the % of GerS pulled-down in each biological replicate. Statistical significance relative to the wild-type complex was determined using one-way ANOVA and Tukey’s test. **, p< 0.005; **** p < 0.0001.