Table 1.

Amino acid changes in the HEHEAA-binding pocket of AapF_GM79 affect HEHEAA binding and protein thermal stability

AapFGM79 seq*	Relative pipA-gfp†	ΔTm +/− HEHEAA‡
None	0.0 (0.4)	NA§
Wild-type	100.0	14
N49P	0.7 (0.4)	0
W389A	1.4 (0.3)	1
Y403A	9.7 (3.7)	1
W404A	1.3 (0.7)	1
T405A	64.0 (14.4)	5
T405P	2.5 (1.4)	ND¶
D407A	16.3 (1.7)	4

^*Amino acid sequence of the indicated Pseudomonas GM79 AapF polypeptides: wild-type, the aapF deletion mutant (none), or the indicated point mutants.

^†HEHEAA activation of the plasmid-borne pipA-gfp reporter in GM79 with the indicated AapF (aapF alleles and pipR are chromosomal). Data are normalized to the wild-type and are means of four biological replicates with the SDs in parentheses. Raw data and complementation data are presented in SI Appendix, Fig. S5.

^‡Melting temperature changes (ΔT_m) (Materials and Methods) of the indicated purified proteins in the presence (+) or absence (−) of 100 μM HEHEAA. Representative thermal shift data are shown in SI Appendix, Fig. S6.

^§Not applicable.

^¶Not determined, no variant protein was obtained.