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. 2014 Mar 3;111(11):4055–4060. doi: 10.1073/pnas.1323285111

Fig. 5.

Fig. 5.

Guanine nucleotide-binding affinity. (A) BODIPY-FL-GTPγS–binding affinity to RocWT was measured using FP assay. Data represented an average of triplicates with error bars (SEM) shown. The dissociation constant (KD) was determined by fitting nonlinear curve regression using GraphPad Prism 6. (B) Competition of prebound BODIPY-labeled GTPγS titrated with natural unlabeled GTP showed that the BODIPY-labeled nucleotide binds to the same site on RocWT as natural nucleotide. (C) BODIPY-FL- GTPγS–binding affinity to RocR1441H was measured using FP assay. Data represented an average of triplicates with error bars (SEM) shown. The dissociation constant was determined as above. (D) BODIPY-FL-GDP–binding affinity to RocWT and RocR1441H was measured using FP assay, and the dissociation constant was determined as above.