TMG-chitotriomycin, an Enzyme Inhibitor Specific for Insect and Fungal β-N-Acetylglucosaminidases, Produced by Actinomycete Streptomyces anulatus NBRC 13369

Abstract

A novel β-N-acetylglucosaminidase (GlcNAcase) inhibitor named TMG-chitotriomycin (1) was isolated from the culture filtrate of Streptomyces anulatus NBRC13369. The strain produced 1 only when colloidal chitin was used as the sole carbon source in the production medium. The structure of 1 was determined by spectral and constitutive sugar analyses of the corresponding alditol derivatives to be an equilibrated mixture of R-D-N,N,N-triMeGlcNH2-(1,4)-β-D-GlcNAc-(1,4)-β-D-GlcNAc-(1,4)-D-GlcNAc and its C-2 epimer of the reducing end residue. TMG-chitotriomycin (1) showed potent and selective inhibition of insect and fungal GlcNAcases with no inhibition of mammalian and plant GlcNAcases. In contrast, the known GlcNAcase inhibitor nagstatin potently inhibited all GlcNAcases. It should be emphasized that synthesized D-N,N,N-triMeGlcNH2, which is the component sugar of 1, showed no inhibition of the insect Spodoptera litura GlcNAcase. These results suggest that the (GlcNAc)3 unit positioned at the reducing end of 1 is essential for its enzyme inhibitory activity. The unique inhibitory spectrum of 1 will be useful to study chitinolytic systems and to develop selective fungicides or pesticides.

Crystal Structures of a Glycoside Hydrolase Family 20 Lacto-N-biosidase from Bifidobacterium bifidum

“…The GlcNAc moieties of these substrates bound to the Ϫ1 subsite were all in conformations close to a 1,4 B conformation (229°Ͻ Ͻ 254°and Ͼ 66°; ideally ϭ 240°and ϭ 90°). Moreover, the complex structure of TMG-chitotriomycin, a linear tetrasaccharide inhibitor (55), bound to the ␤-HexNAcase from Ostrinia furnacalis was reported (30) with the N,N,Ntrimethyl-D-glucosamine group at the nonreducing end of TMG-chitotriomycin also adopting a 1,4 B-like conformation ( ϭ 244.2°and ϭ 84.5°). Therefore, the substrate of the Michaelis complex for GH20 enzymes is thought to adopt a 1,4 B conformation.…”

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

“…To validate the hypothesis, TMG-chitotriomycin [16] was chosen as the starting inhibitor, from which N,N,N-trimethyl-dglucosamine (TMG) derivatives with one to four b-(1!4)-linked GlcNAc residues were designed and synthesized. TMG-chitotriomycin is a recently disclosed and efficient inhibitor for bacterial, fungal, and insect b-N-acetyl-d-hexosaminidases, but has no effect on the enzymes from plants and mammals, [16] a promising feature that might allow for the development of novel and eco-friendly fungicides as well as insecticides.…”

“…TMG-chitotriomycin is a recently disclosed and efficient inhibitor for bacterial, fungal, and insect b-N-acetyl-d-hexosaminidases, but has no effect on the enzymes from plants and mammals, [16] a promising feature that might allow for the development of novel and eco-friendly fungicides as well as insecticides. It is also a pseudotetrasaccharide containing three b-(1!4)-linked GlcNAc units linked to TMG at the nonreducing end.…”

“…Although the structure of TMG-chitotriomycin is known, its inhibition mechanism remains to be clarified. It has been speculated that the charged nitrogen atom in TMG is vital for its inhibitory activity, [16] but Kanzaki et al have shown that TMG itself is inactive against b-N-acetyl-d-hexosaminidase from the insect Spodoptera litura. [16] This surprising result led them to presume that the (GlcNAc) 3 moiety is important for inhibition of b-Nacetyl-d-hexosaminidase.…”

“…It has been speculated that the charged nitrogen atom in TMG is vital for its inhibitory activity, [16] but Kanzaki et al have shown that TMG itself is inactive against b-N-acetyl-d-hexosaminidase from the insect Spodoptera litura. [16] This surprising result led them to presume that the (GlcNAc) 3 moiety is important for inhibition of b-Nacetyl-d-hexosaminidase. The question arising from this is why and how the sugar component affects the interaction between TMG-chitotriomycin and its target enzymes.…”