Literature summary for 4.1.3.4 extracted from

Fu, Z.; Runquist, J.A.; Forouhar, F.; Hussain, M.; Hunt, J.F.; Miziorko, H.M.; Kim, J.J.
Crystal structure of human HMG-CoA lyase: Insights into catalysis and the molecular basis for hydroxymethylglutaric aciduria (2005), J. Biol. Chem., 281, 7526-7532.

Cloned(Commentary)

Cloned (Comment)	Organism
expressed in Escherichia coli	Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
crystal structure at 2.1 A resolution of the recombinant mitochondrial HMG-CoA lyase containing a bound activator cation and 3-hydroxyglutarate. For crystallization experiments, the enzyme is diluted to 5 6 mg/ml. The competitive inhibitor hydroxyglutaryl-CoA is added to the diluted protein at about 1mM concentration. The inhibitor is necessary for generation of uniform diffraction quality crystals. Crystals sufficient for X-ray studies are obtained using an equilibration buffer of 0.1 M Hepes, pH 7.5, 60 mM MgCl2,and 15% polyethylene glycol 8K. The enzyme is mixed 1:1 with the equilibration buffer using sitting drop trays at 19°C. Crystals belong to the monoclinic space group C2 with unit cell parameters a = 197.0 A, b = 117.1 A, c = 86.8 A, and beta = 112.5°. Six monomers are found in the asymmetric unit	Homo sapiens
sitting drop vapour diffusion method with 0.1 M HEPES, pH 7.5, 60 mM MgCl2, and 15% polyethylene glycol 8K	Homo sapiens

Crystallization (Comment)

Organism

crystal structure at 2.1 A resolution of the recombinant mitochondrial HMG-CoA lyase containing a bound activator cation and 3-hydroxyglutarate. For crystallization experiments, the enzyme is diluted to 5 6 mg/ml. The competitive inhibitor hydroxyglutaryl-CoA is added to the diluted protein at about 1mM concentration. The inhibitor is necessary for generation of uniform diffraction quality crystals. Crystals sufficient for X-ray studies are obtained using an equilibration buffer of 0.1 M Hepes, pH 7.5, 60 mM MgCl2,and 15% polyethylene glycol 8K. The enzyme is mixed 1:1 with the equilibration buffer using sitting drop trays at 19°C. Crystals belong to the monoclinic space group C2 with unit cell parameters a = 197.0 A, b = 117.1 A, c = 86.8 A, and beta = 112.5°. Six monomers are found in the asymmetric unit

Homo sapiens

sitting drop vapour diffusion method with 0.1 M HEPES, pH 7.5, 60 mM MgCl2, and 15% polyethylene glycol 8K

Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
D204N	decreased activity	Homo sapiens
D42A	exhibits catalytic rates diminished by 130000fold	Homo sapiens
D42H	decreased activity	Homo sapiens
E279K	unstable recombinant protein	Homo sapiens
H233A	exhibits catalytic rates diminished by 6400fold	Homo sapiens
H233R	decreased activity	Homo sapiens
R41Q	decreased activity	Homo sapiens
S201Y	no activity	Homo sapiens
S75R	no activity	Homo sapiens

Inhibitors

Inhibitors	Comment	Organism	Structure
3-hydroxyglutaryl-CoA	competitive; competitive inhibitor	Homo sapiens

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
mitochondrion	-	Homo sapiens	5739	-

Metals/Ions

Metals/Ions	Comment	Organism	Structure
additional information	identification of His233, His235, ASp42, and Asn275 as metal-binding ligands	Homo sapiens

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
32000	-	SDS-PAGE	Homo sapiens

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	-	-	-

Purification (Commentary)

Purification (Comment)	Organism
Q-Sepharose column chromatography, phenyl-agarose column chromatography, and Superose 12 column chromatography	Homo sapiens

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
(S)-3-Hydroxy-3-methylglutaryl-CoA	-	Homo sapiens	Acetyl-CoA + acetoacetate	-	?

Subunits

Subunits	Comment	Organism
hexamer	X-ray crystallography, the hexamer consists of three dimers which are supposed to be the physiological enzyme form in solution	Homo sapiens

Synonyms

Synonyms	Comment	Organism
3-hydroxy-3-methylglutarate-CoA lyase	-	Homo sapiens
HMG-CoA lyase	-	Homo sapiens