Literature summary for 3.2.1.52 extracted from

Liu, T.; Zhang, H.; Liu, F.; Wu, Q.; Shen, X.; Yang, Q.
Structural determinants of an insect beta-N-Acetyl-D-hexosaminidase specialized as a chitinolytic enzyme (2011), J. Biol. Chem., 286, 4049-4058.

Cloned(Commentary)

Cloned (Comment)	Organism
expression in Pichia pastoris	Ostrinia furnacalis

Crystallization (Commentary)

Crystallization (Comment)	Organism
native enzyme andd in complex with TMG-chitotriomycin, both to 2.1 A resolution. Enzyme displays large conformational changes linked to an open-close mechanism at the entrance of the active site, which is characterized by the lid residue, Trp448. Structure shows a homodimeric enzyme with the two monomers in the adjacent asymmetry units. Each monomer is N-glycosylated at Asn164 and Asn375. All of the 12 cysteine residues of each monomer contribute to the intradisulfide bonds	Ostrinia furnacalis

Crystallization (Comment)

Organism

native enzyme andd in complex with TMG-chitotriomycin, both to 2.1 A resolution. Enzyme displays large conformational changes linked to an open-close mechanism at the entrance of the active site, which is characterized by the lid residue, Trp448. Structure shows a homodimeric enzyme with the two monomers in the adjacent asymmetry units. Each monomer is N-glycosylated at Asn164 and Asn375. All of the 12 cysteine residues of each monomer contribute to the intradisulfide bonds

Ostrinia furnacalis

Protein Variants

Protein Variants	Comment	Organism
E328A	activity similar to wild-type	Ostrinia furnacalis
E328Q	about 20% decrease in activity	Ostrinia furnacalis
H433A	about 20% decrease in activity	Ostrinia furnacalis
V327G	about 30% decrease in activity	Ostrinia furnacalis
W448A	mutation results in a more than 1000fold loss in enzyme activity, due mainly to the effect on kcat	Ostrinia furnacalis
W448F	about 50% decrease in activity	Ostrinia furnacalis
W490A	mutation leads to a 2277fold decrease in sensitivity toward TMG-chitotriomycin as well as an 18fold decrease in binding affinity for the substrate (GlcNAc)2	Ostrinia furnacalis

Inhibitors

Inhibitors	Comment	Organism	Structure
TMG-chitotriomycin	mechanism of selective inhibition is related to the existence of the +1 subsite at the active pocket of OfHex1 and a key residue, Trp490, at this site. Mutation of Trp490 to Ala leads to a 2277fold decrease in sensitivity toward TMG-chitotriomycin as well as an 18-fold decrease in binding affinity for the substrate (GlcNAc)2	Ostrinia furnacalis

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
0.07	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant V327G, pH 7.0, 25°C	Ostrinia furnacalis
0.095	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant H433A, pH 7.0, 25°C	Ostrinia furnacalis
0.096	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant E328A, pH 7.0, 25°C	Ostrinia furnacalis
0.1	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant W490A, pH 7.0, 25°C	Ostrinia furnacalis
0.105	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant E328Q, pH 7.0, 25°C	Ostrinia furnacalis
0.107	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	wild-type, pH 7.0, 25°C	Ostrinia furnacalis
0.126	-	chitobiose	mutant V327G, pH 7.0, 25°C	Ostrinia furnacalis
0.148	-	chitobiose	wild-type, pH 7.0, 25°C	Ostrinia furnacalis
0.149	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant W448F, pH 7.0, 25°C	Ostrinia furnacalis
0.223	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant W448A, pH 7.0, 25°C	Ostrinia furnacalis
1.875	-	chitobiose	mutant W490A, pH 7.0, 25°C	Ostrinia furnacalis

Organism

Organism	UniProt	Comment	Textmining
Ostrinia furnacalis	Q06GJ0	-	-

Posttranslational Modification

Posttranslational Modification	Comment	Organism
glycoprotein	each monomer is N-glycosylated at Asn164 and Asn375	Ostrinia furnacalis

Source Tissue

Source Tissue	Comment	Organism	Textmining
pupa	up-regulation of OfHEX1 transcription, specific to the integument during the pupation	Ostrinia furnacalis	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
4-methylumbelliferyl N-acetyl-beta-D-glucosaminide + H2O	-	Ostrinia furnacalis	N-acetyl-beta-D-glucosamine + 4-methylumbelliferone	-	?
chitobiose + H2O	-	Ostrinia furnacalis	2 D-glucosamine	-	?

Subunits

Subunits	Comment	Organism
dimer	data from crystal structure	Ostrinia furnacalis

Synonyms

Synonyms	Comment	Organism
Hex1	-	Ostrinia furnacalis

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism
0.31	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant H433A, pH 7.0, 25°C	Ostrinia furnacalis
0.47	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant W448A, pH 7.0, 25°C	Ostrinia furnacalis
0.57	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant W448F, pH 7.0, 25°C	Ostrinia furnacalis
168	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant W490A, pH 7.0, 25°C	Ostrinia furnacalis
189	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant V327G, pH 7.0, 25°C	Ostrinia furnacalis
337	-	chitobiose	mutant V327G, pH 7.0, 25°C	Ostrinia furnacalis
349	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant E328Q, pH 7.0, 25°C	Ostrinia furnacalis
356	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant E328A, pH 7.0, 25°C	Ostrinia furnacalis
434.7	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	wild-type, pH 7.0, 25°C	Ostrinia furnacalis
450	-	chitobiose	mutant W490A, pH 7.0, 25°C	Ostrinia furnacalis
507	-	chitobiose	wild-type, pH 7.0, 25°C	Ostrinia furnacalis

General Information

General Information	Comment	Organism
physiological function	knock-down by RNAi. Before pupation, most of the larvae injected with RNA1 survive without any visible changes in phenotype and appear similar to those injected with control. At the pupation stage, 20% of the RNAi-injected larvae show abnormal phenotypes that are absent in control larvae and die 1 or 2 days after pupation. All of the larvae with abnormal phenotypes fail to shed their old cuticles completely before new ones started to form underneath	Ostrinia furnacalis

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism
2.1	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant W448A, pH 7.0, 25°C	Ostrinia furnacalis
3.3	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant H433A, pH 7.0, 25°C	Ostrinia furnacalis
3.8	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant W448F, pH 7.0, 25°C	Ostrinia furnacalis
240	-	chitobiose	mutant W490A, pH 7.0, 25°C	Ostrinia furnacalis
1676	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant W490A, pH 7.0, 25°C	Ostrinia furnacalis
2675	-	chitobiose	mutant V327G, pH 7.0, 25°C	Ostrinia furnacalis
2700	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant V327G, pH 7.0, 25°C	Ostrinia furnacalis
3324	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant E328Q, pH 7.0, 25°C	Ostrinia furnacalis
3428	-	chitobiose	wild-type, pH 7.0, 25°C	Ostrinia furnacalis
3708	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	mutant E328A, pH 7.0, 25°C	Ostrinia furnacalis
4063	-	4-methylumbelliferyl N-acetyl-beta-D-glucosaminide	wild-type, pH 7.0, 25°C	Ostrinia furnacalis