Phosphoramidase activity of the E. coli hinT by 31 P NMR Assay. (A) E. coli hinT protein (2.5μg) incubated with AMP-Mor (10mM) at 22°C for 36min. (B) for 66min.
Hint is required for growth on the LB-agar media containing 0.75, KCl, 0.9 M NaCl, 0.5 M NaOAc, and 10mM MnCl2. The E. coli hinT knockout strain BB3 (hinTΔ) was transformed with either an empty vector (hinTΔ pACYC), a plasmid expressing E.coli hinT (hinTΔ pACYC-hinT), or a plasmid expressing the His-101 to Ala mutant (hinTΔ pACYC-hinT-H101A). Colonies were observed for BB3 cells transformed with pACYC-hinT (hinTΔ pACYC-hinT) and pACYC-hinT-H101A (hinTΔ, pACYC-hinT-H101A) when grown with LB-agar media containing 0.75M KCl, 0.9 M NaCl, and 10mM MnCl2. When cultures were grown in the presence of 0.5 M NaOAc, colonies were observed for BB3 cells transformed with pACYC-hinT, while a marked reduction in the number of colonies for cells transformed with pACYC-hinT-H101A could be seen.