Literature summary for 3.2.1.129 extracted from

Schwarzer, D.; Stummeyer, K.; Haselhorst, T.; Freiberger, F.; Rode, B.; Grove, M.; Scheper, T.; von Itzstein, M.; Muehlenhoff, M.; Gerardy-Schahn, R.
Proteolytic release of the intramolecular chaperone domain confers processivity to endosialidase f (2009), J. Biol. Chem., 284, 9465-9474.

Search for more literature for 3.2.1.129

Cloned(Commentary)

Cloned (Comment)	Organism
protein expression in Escherichia coli BL21-Gold(DE3) in the presence of 100 microgram/ml Carbenicillin, 30°C	Escherichia phage K1F

Crystallization (Commentary)

Crystallization (Comment)	Organism
crystal structure of the catalytic domain of endoN from caliphate K1F reveals a functional trimer, folding is mediated by an intramolecular C-terminal chaperone domain	Escherichia phage K1F

Protein Variants

Protein Variants	Comment	Organism
Q853A	binding site mutation, active within control range with soluble polysialic acid	Escherichia phage K1F
R596A/R647A	control active site mutation without affecting maturation or binding, EC50: 1.9 nM polysialic acid	Escherichia phage K1F
R596A/R647A/Q853A	active site mutation plus binding site mutation, EC50: 5.0 nM surface bound polysialic acid	Escherichia phage K1F
R596A/R647A/R837A	active site mutation plus binding site mutation, 5-fold increased EC50: 11 nM polysialic acid	Escherichia phage K1F
R596A/R647A/R837A/S848A	active site mutation plus binding site mutation, EC50: 30 nM surface bound polysialic acid, increased EC50: 30 nM polysialic acid	Escherichia phage K1F
R596A/R647A/S848A	active site mutation plus binding site mutation, only binding site mutant with SDS resistance which is a criterion for kinetic stabilization of the enzyme, EC50: 4.1 nM surface bound polysialic acid	Escherichia phage K1F
R596A/R647A/S848A/Q853A	active site mutation plus binding site mutation, increased EC50: 6.2 nM polysialic acid	Escherichia phage K1F
R596A/R647A/S911A	active site mutation plus cleavage site mutation, tremendously increased EC50: 360 nM polysialic acid	Escherichia phage K1F
R837A	binding site mutation, increased molar activity with soluble polysialic acid	Escherichia phage K1F
R837A/Q853A	binding site mutation, insoluble enzyme	Escherichia phage K1F
R837A/S848A	binding site mutation, increased molar activity with soluble polysialic acid	Escherichia phage K1F
R837A/S848A/Q853A	binding site mutation, insoluble enzyme	Escherichia phage K1F
S848A	binding site mutation, active within control range with soluble polysialic acid	Escherichia phage K1F
S848A/Q853A	binding site mutation, active within control range with soluble polysialic acid	Escherichia phage K1F
S911A	cleavage site mutation: prevents proteolysis of the C-terminal domain that functions as an intramolecular chaperone and is normally released during enzyme maturation. Substrate binding is reduced similarly to binding site mutations. Altered activities: 3times higher activity with soluble polycialic acid as substrate, 190fold reduced activity with immobilized polysialic acid as substrate, no difference in activity with minimal substrate tetrameric sialic acid	Escherichia phage K1F

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
0.85	-	tetrameric silica acid	identical kinetic parameters for wild type and cleavage site mutant S911A	Escherichia phage K1F

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
polysialic acid capsules of bacteria + H2O	Escherichia phage K1F	minimum substrate is tetrameric polysialic acid, processive enzyme activity on oligomers larger than that, confirmation by H NMR spectroscopy and anion-exchange chromatography	oligomers of alpha 2,8-linked polysialic acid	-	?

Organism

Organism	UniProt	Comment	Textmining
Escherichia phage K1F	-	-	-

Posttranslational Modification

Posttranslational Modification	Comment	Organism
proteolytic modification	proteolysis of the C-terminal domain that functions as an intramolecular chaperone and is released during enzyme maturation of wild-type enzyme	Escherichia phage K1F

Purification (Commentary)

Purification (Comment)	Organism
description in Schwarzer, D. et al. (2007) J.Biol. Chem. 282, 2821-2831	Escherichia phage K1F

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
longchain alpha 2,8-linked polysialic acid + H2O	minimum substrate is tetrameric polysialic acid, processive enzyme activity on oligomers larger than that, confirmation by H NMR spectroscopy and anion-exchange chromatography	Escherichia phage K1F	oligomers of alpha 2,8-linked polysialic acid	major product is an oligomer consisting of 3 monomers in wild-type and cleavage mutant S911A, in binding site mutant R837A/S848A more random oligomers are produced	?
polysialic acid capsules of bacteria + H2O	minimum substrate is tetrameric polysialic acid, processive enzyme activity on oligomers larger than that, confirmation by H NMR spectroscopy and anion-exchange chromatography	Escherichia phage K1F	oligomers of alpha 2,8-linked polysialic acid	-	?
tetrameric silica acid + H2O	-	Escherichia phage K1F	?	-	?

Synonyms

Synonyms	Comment	Organism
endo-N-acetylneuraminidase	-	Escherichia phage K1F
endoNF	from coliphage K1F, lacking N-terminal capsid binding domain	Escherichia phage K1F
endosialidase	-	Escherichia phage K1F

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
4.37	-	tetrameric silica acid	identical kinetic parameters for wild type and cleavage site mutant S911A	Escherichia phage K1F

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Release 2023.1 (January 2023)