Communication
ChemComm
24 towards the Orn-activating domain of GrsB are attributed to
This work was partly supported by a Grant-in Aid for Young
the common electrostatic interactions between the carboxylate Scientists (B) 26750370 (F.I.) and research grants from the
of Glu278 and the side chain functionalities of the compounds. Japan Society of the Promotion of Science and the Ministry of
Since the Orn-activating domain of HSAF synthetase can activate Education, Culture, Sports, Science and Technology in Japan
both L-Orn and L-Lys among the 16 amino acids tested,27 it is (MEXT) (H.K.). We thank Prof. Mohamed Marahiel (Philipps-
¨
postulated that the Orn-activating domain of GrsB could accept Universitat Marburg, Germany) for providing the GrsA and
L-Gln, L-Lys and L-Arg as alternative substrates.
TycB1 expression constructs.
To evaluate the Val- and Leu-activating domains of GrsB in
competitive experiments, the DSM 5759 proteome was individually
treated with 100 mM of each aminoacyl-AMP analog 6–25 before the
addition of probes 4 and 5. Assessment of the inhibition potency
revealed that five (12, 13, 18, 20 and 25) and three (10, 12 and 18)
of the 20 compounds displayed high (Z90%) inhibition of the
Val- and Leu-activating domains of GrsB, respectively (Fig. 4c and d,
Fig. S20 and S22, ESI†). Inhibitors 12, 13, 18, 20 and 25 gave IC50
values of 2.8 ꢀ 0.23 mM, 6.0 ꢀ 0.10 mM, 0.42 ꢀ 0.08 mM, 0.65 ꢀ
0.04 mM and 2.2 ꢀ 0.28 mM for the inhibition of the Val-activating
domain of GrsB (Fig. S21 and Table S6, ESI†). A dose–response
curve by the probe’s cognate competitor 11 afforded an IC50 value
of 0.11 ꢀ 0.04 mM (Fig. S21 and Table S6, ESI†). Dose–response
curves for the inhibition of the Leu-activating domain of GrsB by 12
and 18 gave IC50 values of 1.7 ꢀ 0.09 nM and 2.7 ꢀ 0.22 mM,
respectively (Fig. S23 and Table S7, ESI†). In contrast, 10 did not
provide an inhibitor sensitive profile towards the Leu-activating
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