Literature summary for 3.7.1.8 extracted from

Lack, N.; Yam, K.; Lowe, E.; Horsman, G.; Owen, R.; Sim, E.; Eltis, L.
Characterization of a carbon-carbon hydrolase from Mycobacterium tuberculosis involved in cholesterol metabolism (2010), J. Biol. Chem., 285, 434-443.

Crystallization (Commentary)

Crystallization (Comment)	Organism
mutant S114A in complex with 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid, 8-(2-chlorophenyl)-2-hydroxy-5-methyl-6-oxoocta-2,4-dienoic acid, or 4,5-9,10-diseco-3-hydroxy-5,9,17-trioxoandrosta-1(10),2-diene-4-oic acid, sitting drop vapor diffusion, 0.001 ml of 8 mg/ml protein solution is mixed with 0.001 ml of precipitant solution containing 200 mM KSCN, 24% PEG 3350, and 100 mM bis-tris propane, pH 7.0, soaking of S114A crystals in 0.01 ml of precipitant solution supplemented with 15 mM of each ligand for 30-60 min, X-ray diffractin structure determination and analysis at 1.8-2.1 A resolution, molecular replacement	Mycobacterium tuberculosis

Crystallization (Comment)

Organism

mutant S114A in complex with 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid, 8-(2-chlorophenyl)-2-hydroxy-5-methyl-6-oxoocta-2,4-dienoic acid, or 4,5-9,10-diseco-3-hydroxy-5,9,17-trioxoandrosta-1(10),2-diene-4-oic acid, sitting drop vapor diffusion, 0.001 ml of 8 mg/ml protein solution is mixed with 0.001 ml of precipitant solution containing 200 mM KSCN, 24% PEG 3350, and 100 mM bis-tris propane, pH 7.0, soaking of S114A crystals in 0.01 ml of precipitant solution supplemented with 15 mM of each ligand for 30-60 min, X-ray diffractin structure determination and analysis at 1.8-2.1 A resolution, molecular replacement

Mycobacterium tuberculosis

Protein Variants

Protein Variants	Comment	Organism
S114A	active site HsaD mutant, catalytically impaired and binds 4,5-9,10-diseco-3-hydroxy-5,9,17-trioxoandrosta-1(10),2-diene-4-oic acid with an altered dissociation constant compared to the wild-type enzyme	Mycobacterium tuberculosis

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
0.008	-	2-Hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid	pH 7.5, 25°C	Mycobacterium tuberculosis
0.31	-	8-(2-chlorophenyl)-2-hydroxy-5-methyl-6-oxoocta-2,4-dienoic acid	pH 7.5, 25°C	Mycobacterium tuberculosis

Organism

Organism	UniProt	Comment	Textmining
Mycobacterium tuberculosis	P9WNH5	-	-
Mycobacterium tuberculosis H37Rv	P9WNH5	-	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid + H2O	i.e. HOPDA, a biphenyl meta-cleavage product	Mycobacterium tuberculosis	2-hydroxypenta-2,4-dienoic acid + benzoate	-	?
2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid + H2O	i.e. HOPDA, a biphenyl meta-cleavage product	Mycobacterium tuberculosis H37Rv	2-hydroxypenta-2,4-dienoic acid + benzoate	-	?
4,5-9,10-diseco-3-hydroxy-5,9,17-trioxoandrosta-1(10),2-diene-4-oic acid + H2O	i.e. DSHA, a cholesterol meta-cleavage product	Mycobacterium tuberculosis	3-[(7aR)-7a-hydroxy-1,5-dioxooctahydro-1H-inden-4-yl]propanoic acid + (2Z,4Z)-5-hydroxyhexa-2,4-dienoic acid	-	?
4,5-9,10-diseco-3-hydroxy-5,9,17-trioxoandrosta-1(10),2-diene-4-oic acid + H2O	i.e. DSHA, a cholesterol meta-cleavage product	Mycobacterium tuberculosis H37Rv	3-[(7aR)-7a-hydroxy-1,5-dioxooctahydro-1H-inden-4-yl]propanoic acid + (2Z,4Z)-5-hydroxyhexa-2,4-dienoic acid	-	?
8-(2-chlorophenyl)-2-hydroxy-5-methyl-6-oxoocta-2,4-dienoic acid + H2O	i.e. HOPODA, a HOPDA synthetic analogue	Mycobacterium tuberculosis	2-hydroxypenta-2,4-dienoic acid + benzoate	-	?
8-(2-chlorophenyl)-2-hydroxy-5-methyl-6-oxoocta-2,4-dienoic acid + H2O	i.e. HOPODA, a HOPDA synthetic analogue	Mycobacterium tuberculosis H37Rv	2-hydroxypenta-2,4-dienoic acid + benzoate	-	?

Synonyms

Synonyms	Comment	Organism
2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate hydrolase	-	Mycobacterium tuberculosis
HsaD	-	Mycobacterium tuberculosis

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
25	-	assay at	Mycobacterium tuberculosis

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
0.066	-	2-Hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid	pH 7.5, 25°C	Mycobacterium tuberculosis
0.33	-	8-(2-chlorophenyl)-2-hydroxy-5-methyl-6-oxoocta-2,4-dienoic acid	pH 7.5, 25°C	Mycobacterium tuberculosis

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Mycobacterium tuberculosis

General Information

General Information	Comment	Organism
physiological function	the enzyme is involved in the intracellular survival of pathogen Mycobacterium tuberculosis	Mycobacterium tuberculosis

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism	Structure
1.1	-	8-(2-chlorophenyl)-2-hydroxy-5-methyl-6-oxoocta-2,4-dienoic acid	pH 7.5, 25°C	Mycobacterium tuberculosis
9	-	2-Hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid	pH 7.5, 25°C	Mycobacterium tuberculosis