Literature summary for 2.5.1.61 extracted from

Clavero, S.; Bishop, D.F.; Haskins, M.E.; Giger, U.; Kauppinen, R.; Desnick, R.J.
Feline acute intermittent porphyria: a phenocopy masquerading as an erythropoietic porphyria due to dominant and recessive hydroxymethylbilane synthase mutations (2010), Hum. Mol. Genet., 19, 584-596.

Application

Application	Comment	Organism
medicine	cats with naturally occurring mutant enzymes can be used as animal model for acute intermittent porphyria	Felis catus

Cloned(Commentary)

Cloned (Comment)	Organism
expression in Escherichia coli BL21+	Felis catus

Protein Variants

Protein Variants	Comment	Organism
A84T	type OregonAR (c250G>A), autosomal recessive, therefore phenotype is homozygous, about 35% wild type activity	Felis catus
L64S	c.189dupT mutant type Saskatchewan with duplication that leads to a frameshift and thus to a trunkated enzyme due to a premature stop codon 65 and the exchange L64S, autosomal dominant	Felis catus
additional information	4 distinct mutations cause acute intermittent porphyria in cats, mutant phenotype has brownish discolored teeth and brownish urine, both fluorescent under UV light, accumulation of substrates, mutations affect housekeeping and erythroid-specific isozymes in the same way	Felis catus
additional information	4 distinct mutations cause acute intermittent porphyria in cats, mutant phenotype has brownish discolored teeth and brownish urine, both fluorescent under UV light, mutations affect housekeeping and erythroid-specific isozymes in the same way	Felis catus
additional information	type Massachusetts, in-frame 3 bp deletion (c.842_844delGAG = delGly281), autosomal dominant, less than 1% wild type activity	Felis catus
R149W	type Missouris (c.445C>T), substitution identical to human mutation but there leading to an nonsense mutation, autosomal dominant, less than 1% wild type activity	Felis catus

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
40500	-	erythroid enzyme, SDS-PAGE	Felis catus
43000	-	housekeeping enzyme, SDS-PAGE	Felis catus

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
4 porphobilinogen + H2O	Felis catus	-	hydroxymethylbilane + 4 NH3	-	?

Organism

Organism	UniProt	Comment	Textmining
Felis catus	D3W810	wild type siblings of mutant cats or unrelated wild type cats, porphyric mutant cats type Saskatchewan (c.189dupT = trunkated (premature stop codon 65) L64S), type Massachusetts (c842_844delGAG = delGly281), type Missouris (c.445C>T = R149W), type OregonAR (c250G>A = A84T)	-
Felis catus	D3W811	wild type siblings of mutant cats or unrelated wild type cats, porphyric mutant cats type Saskatchewan (c.189dupT = trunkated (premature stop codon 65) L64S), type Massachusetts (c842_844delGAG = delGly281), type Missouris (c.445C>T = R149W), type OregonAR (c250G>A = A84T)	-
Felis catus	D3W812	wild type siblings of mutant cats or unrelated wild type cats, porphyric mutant cats type Saskatchewan (c.189dupT = trunkated (premature stop codon 65) L64S), type Massachusetts (c842_844delGAG = delGly281), type Missouris (c.445C>T = R149W), type OregonAR (c250G>A = A84T)	-

Purification (Commentary)

Purification (Comment)	Organism
packed red blood cells are washed in phosphate buffered saline, lysed with buffer containing 0.1 M KHEPES, pH 7.5, 0.1% Triton X-100, 1 mM DTT, and 0.02% sodium azide, liver and spleen are diced and homogenized in buffer containing 50 mM KHEPES, pH 7.5, 150 mM KCl, 1 mM Na2EDTA, 1 mM DTT, 0.1 mM PMSF, for activity and thermostability experiments enzymes are expressed in Escherichia coli, cells are harvested and resuspended in 50 mM HEPES, pH 7.8, 0.5 M NaCl, 5 mM DTT, 330 microg/ml lysozyme, 5 microg/ml DNAse and RNAse, freeze-thawed 3times in dry-ice/ethanol, centrifuged, adjusted to pH 7.4 and 8.0 with NaOH, incubated at 37°C or 50°C for 1 h	Felis catus

Purification (Comment)

Organism

packed red blood cells are washed in phosphate buffered saline, lysed with buffer containing 0.1 M KHEPES, pH 7.5, 0.1% Triton X-100, 1 mM DTT, and 0.02% sodium azide, liver and spleen are diced and homogenized in buffer containing 50 mM KHEPES, pH 7.5, 150 mM KCl, 1 mM Na2EDTA, 1 mM DTT, 0.1 mM PMSF, for activity and thermostability experiments enzymes are expressed in Escherichia coli, cells are harvested and resuspended in 50 mM HEPES, pH 7.8, 0.5 M NaCl, 5 mM DTT, 330 microg/ml lysozyme, 5 microg/ml DNAse and RNAse, freeze-thawed 3times in dry-ice/ethanol, centrifuged, adjusted to pH 7.4 and 8.0 with NaOH, incubated at 37°C or 50°C for 1 h

Felis catus

Source Tissue

Source Tissue	Comment	Organism	Textmining
erythrocyte	-	Felis catus	-
liver	-	Felis catus	-
spleen	-	Felis catus	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
4 porphobilinogen + H2O	-	Felis catus	hydroxymethylbilane + 4 NH3	-	?

Synonyms

Synonyms	Comment	Organism
HMBS1a-synthase	housekeeping, GenBank GQ850461, isolated from liver	Felis catus
HMBS1b-synthase	housekeeping, truncated form, GenBank GQ850462, isolated from liver	Felis catus
HMBS2a-synthase	erythroid-specific, GenBank GQ850463, isolated from spleen	Felis catus
HMBS2b-synthase	erythroid-specific, GenBank GQ850464, isolated from spleen	Felis catus
hydroxymethylbilane synthase	-	Felis catus

Temperature Stability [°C]

Temperature Stability Minimum [°C]	Temperature Stability Maximum [°C]	Comment	Organism
50	-	A84T mutant houskeeping enzyme (OregonAR cat) at pH 8.0 has a prolonged half-life of 6.9 h compared to 1.9 h for the wild-type	Felis catus

General Information

General Information	Comment	Organism
malfunction	half-maximal activity of the enzyme causes acute intermittent porphyria due to an error of heme biosynthesis	Felis catus