Literature summary for 3.1.2.6 extracted from

Urscher, M.; Deponte, M.
Plasmodium falciparum glyoxalase II: Theorell-Chance product inhibition patterns, rate-limiting substrate binding via Arg(257)/Lys(260), and unmasking of acid-base catalysis (2009), Biol. Chem., 390, 1171-1183.

Search for more literature for 3.1.2.6

Cloned(Commentary)

Cloned (Comment)	Organism
into pQE30 vector. N-terminally MRGS(H)6GS-tagged GloII wild-type and mutant enzymes transformed into Escherichia coli competent XL1-Blue cells or EZ cells	Plasmodium falciparum

Protein Variants

Protein Variants	Comment	Organism
K260D	unaltered metal binding sites and is correctly folded, but reduced activity. Has a much lower catalytic efficiency, but the spectrum is very similar to the wild-type enzyme. Has a highly simplified bell-shaped pH-activity profile with a maximum at pH 6.7 in contrast to the broad asymmetric profiles of the other enzymes	Plasmodium falciparum
K260Q	unaltered metal binding sites and is correctly folded, but reduced activity	Plasmodium falciparum
R154K	unaltered metal binding sites and is correctly folded, but reduced activity	Plasmodium falciparum
R154M	unaltered metal binding sites and is correctly folded, but reduced activity	Plasmodium falciparum
R257D	unaltered metal binding sites and is correctly folded, but reduced activity	Plasmodium falciparum
R257Q	unaltered metal binding sites and is correctly folded, but reduced activity	Plasmodium falciparum

Inhibitors

Inhibitors	Comment	Organism	Structure
D-lactate	competitive inhibition	Plasmodium falciparum
glutathione	non-competitive inhibition. Is a stronger inhibitor than the ethyl ester of glutathione. Theorell-Chance Bi Bi mechanism with glutathione as the second product	Plasmodium falciparum
glutathione ethyl ester	carboxylate group of the glycine-moiety is modified, non-competitive inhibition	Plasmodium falciparum

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
0.114	-	S-D-lactoylglutathione	wild-type, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
0.116	-	S-D-lactoylglutathione	mutant R154K, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
0.275	-	S-D-lactoylglutathione	mutant R154M, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
2.551	-	S-D-lactoylglutathione	mutant R257Q, at 25°C, in 100 mM MOPS/NaOH, pH 7.4	Plasmodium falciparum
2.867	-	S-D-lactoylglutathione	mutant K260D, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
4.308	-	S-D-lactoylglutathione	mutant K260Q, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
5.782	-	S-D-lactoylglutathione	mutant R257D, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Zinc	recombinant GloII is zinc-dependent	Plasmodium falciparum

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
31800	-	x * 31800, His-tagged GloII, SDS-PAGE	Plasmodium falciparum

Organism

Organism	UniProt	Comment	Textmining
Plasmodium falciparum	-	-	-

Purification (Commentary)

Purification (Comment)	Organism
wild-type and glutathione-binding site mutants, by sonication, centrifugation and on Ni-NTA column	Plasmodium falciparum

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
S-D-lactoylglutathione + H2O	substrate is predominantly bound via ionic interactions with the conserved residues Arg257 and Lys260. Correct substrate binding is a pH- and salt-dependent rate-limiting step for catalysis. Bi Bi mechanism with the direct involvement of a hydroxide ion as a second substrate	Plasmodium falciparum	D-lactate + glutathione	-	?

Subunits

Subunits	Comment	Organism
?	x * 31800, His-tagged GloII, SDS-PAGE	Plasmodium falciparum

Synonyms

Synonyms	Comment	Organism
GloII	-	Plasmodium falciparum
glyoxalase II	-	Plasmodium falciparum

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
60	-	S-D-lactoylglutathione	mutant R257D, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
67	-	S-D-lactoylglutathione	mutant R257Q, at 25°C, in 100 mM MOPS/NaOH, pH 7.4	Plasmodium falciparum
74	-	S-D-lactoylglutathione	mutant K260D, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
171	-	S-D-lactoylglutathione	mutant K260Q, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
222	-	S-D-lactoylglutathione	mutant R154M, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
285	-	S-D-lactoylglutathione	mutant R154K, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
375	-	S-D-lactoylglutathione	wild-type, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum

Ki Value [mM]

Ki Value [mM]	Ki Value maximum [mM]	Inhibitor	Comment	Organism	Structure
0.000031	-	D-lactate	inhibitor binding to the free enzyme, at 25°C, pH 6.8	Plasmodium falciparum
3.5	-	glutathione	inhibitor binding to the free enzyme, at 25°C, pH 6.8	Plasmodium falciparum
13	-	glutathione	inhibitor binding to the occupied enzyme, at 25°C, pH 6.8	Plasmodium falciparum
14	-	glutathione ethyl ester	inhibitor binding to the free enzyme, at 25°C, pH 6.8	Plasmodium falciparum
49	-	glutathione ethyl ester	inhibitor binding to the occupied enzyme, at 25°C, pH 6.8	Plasmodium falciparum

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism	Structure
11	-	S-D-lactoylglutathione	mutant R257D, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
26	-	S-D-lactoylglutathione	mutant K260D, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
30	-	S-D-lactoylglutathione	mutant R257Q, at 25°C, in 100 mM MOPS/NaOH, pH 7.4	Plasmodium falciparum
41	-	S-D-lactoylglutathione	mutant K260Q, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
810	-	S-D-lactoylglutathione	mutant R154M, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
2400	-	S-D-lactoylglutathione	mutant R154K, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum
3300	-	S-D-lactoylglutathione	wild-type, at 25°C, in 100 mM MOPS/NaOH, pH 6.8	Plasmodium falciparum

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Release 2023.1 (January 2023)