Synthesis, Evaluation, and Mechanism of N,N,N-Trimethyl-D-glucosamine-(1→4)-chitooligosaccharides as Selective Inhibitors of Glycosyl Hydrolase Family 20 β-N-Acetyl-D-hexosaminidases

GH20 beta-N-acetyl-D-hexosaminidases are enzymes involved in many vital processes. Inhibitors that specifically target GH20 enzymes in pests are of agricultural and economic importance. Structural comparison has revealed that the bacterial chitindegrading beta-N-acetyl-D-hexosaminidases each have an extra +1 subsite in the active site; this structural difference could be exploited for the development of selective inhibitors. N,N,N-trimethyl-D-glucosamine (TMG)-chitotriomycin, which contains three GlcNAc residues, is a natural selective inhibitor against bacterial and insect beta-N-acetyl-D-hexosaminidases. However, our structural alignment analysis indicated that the two GlcNAc residues at the reducing end might be unnecessary. To prove this hypothesis, we designed and synthesized a series of TMG-chitotriomycin analogues containing one to four GlcNAc units. Inhibitory kinetics and molecular docking showed that TMG-(GlcNAc)(2) , is as active as TMG-chitotriomycin [TMG-(GlcNAc)(3)]. The selective inhibition mechanism of TMG-chitotriomycin was also explained.