Literature summary for 4.1.2.47 extracted from

Lauble, H.; Miehlich, B.; F�rster, S.; Kobler, C.; Wajant, H.; Effenberger, F.
Structure determinants of substrate specificity of hydroxynitrile lyase from Manihot esculenta (2002), Protein Sci., 11, 65-71.

Cloned(Commentary)

Cloned (Comment)	Organism
expression in Escherichia coli	Manihot esculenta

Crystallization (Commentary)

Crystallization (Comment)	Organism
vapor-diffusion hanging-drop method. The crystal structure of the mutant W128A substrate free form at 2.1 A resolution indicates that the W128A substitution has significantly enlarged the active-site channel entrance. The MeHNL-W128A/4-hydroxybenzaldehyde complex structure at 2.1 A resolution shows the presence of two hydroxybenzaldehyde molecules in a sandwich type arrangement in the active site with an additional hydrogen bridge to the reacting center	Manihot esculenta

Crystallization (Comment)

Organism

vapor-diffusion hanging-drop method. The crystal structure of the mutant W128A substrate free form at 2.1 A resolution indicates that the W128A substitution has significantly enlarged the active-site channel entrance. The MeHNL-W128A/4-hydroxybenzaldehyde complex structure at 2.1 A resolution shows the presence of two hydroxybenzaldehyde molecules in a sandwich type arrangement in the active site with an additional hydrogen bridge to the reacting center

Manihot esculenta

Protein Variants

Protein Variants	Comment	Organism
W128A	activity with the natural substrate 2-hydroxy-2-methylpropanenitrile (acetone cyanohydrin) is 70% of wild-type activity. The specific activities of MeHNL-W128A for the unnatural substrates mandelonitrile and 4-hydroxymandelonitrile are increased 9fold and 450fold, respectively, compared with the wild-type. The crystal structure of the mutant W128A substrate free form at 2.1 A resolution indicates that the W128A substitution has significantly enlarged the active-site channel entrance, and thereby explains the observed changes in substrate specificity for bulky substrates	Manihot esculenta

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
additional information	-	additional information	wild-type enzyme shows very low activity with 4-hydroxymandelonitrile, activity of mutant enzyme W128A is increased 450fold compared to wild-type value (Km-value for mutant enzyme W128A is 0.625 mM)	Manihot esculenta
4.5	-	(S)-mandelonitrile	pH 5.2, 23°C, mutant enzyme W128A	Manihot esculenta
30	-	(S)-mandelonitrile	pH 5.2, 23°C, wild-type enzyme	Manihot esculenta
67	-	2-hydroxy-2-methylpropanenitrile	pH 5.2, 23°C, wild-type enzyme	Manihot esculenta
150	-	2-hydroxy-2-methylpropanenitrile	pH 5.2, 23°C, mutant enzyme W128A	Manihot esculenta

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
2-hydroxy-2-methylpropanenitrile	Manihot esculenta	-	acetone + HCN	-	?

Organism

Organism	UniProt	Comment	Textmining
Manihot esculenta	P52705	-	-

Purification (Commentary)

Purification (Comment)	Organism
-	Manihot esculenta

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
(S)-mandelonitrile	-	Manihot esculenta	cyanide + benzaldehyde	-	?
2-hydroxy-2-methylpropanenitrile	-	Manihot esculenta	acetone + HCN	-	?
2-hydroxy-2-methylpropanenitrile	i.e. acetone cyanohydrin	Manihot esculenta	cyanide + acetone	-	?
additional information	wild-type enzyme shows very low activity with 4-hydroxymandelonitrile, activity of mutant enzyme W128A is increased 450fold compared to wild-type value (Km-value for mutant enzyme W128A is 0.625 mM)	Manihot esculenta	?	-	?

Synonyms

Synonyms	Comment	Organism
MeHNL	-	Manihot esculenta

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
23	-	assay at	Manihot esculenta

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
5.2	-	assay at	Manihot esculenta