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Literature summary for 1.14.11.2 extracted from

  • Lamberg, A.; Pihlajaniemi, T.; Kivirikko, K.I.
    Site-directed mutagenesis of the a subunit of human prolyl 4-hydroxylase. Identification of three histidine residues critical for catalytic activity (1995), J. Biol. Chem., 270, 9926-9931.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expression of histidine and cysteine mutant alpha subunits together with the wild-type beta subunit in insect cells by means of baculovirus vectors Homo sapiens

Protein Variants

Protein Variants Comment Organism
C150S the mutation has no major effect on tetramer assembly, but the amount of tetramer is slightly reduced, being about 80% of that of the wild-type enzyme Homo sapiens
C486S the mutation totally prevents tetramer assembly Homo sapiens
C511S the mutation totally prevents tetramer assembly Homo sapiens
H141S the mutation has no effect on enzyme activity and does not inhibit tetramer assembly Homo sapiens
H165S the mutation produces a reduction of about 60% in enzyme activity per unit extractable cell protein relative to that obtained with the wild-type alpha subunit, the amount of tetramer is reduced by about 20-25%, the Km values for Fe2+, 2-oxoglutarate, ascorbate and the peptide substrate with the mutant are identical to those with the wild-type enzyme Homo sapiens
H221S the mutation produces a reduction of about 30% in enzyme activity per unit extractable cell protein relative to that obtained with the wild-type alpha subunit, the amount of tetramer is reduced by about 20-25%, the Km values for Fe2+, 2-oxoglutarate, ascorbate and the peptide substrate with the mutant are identical to those with the wild-type enzyme Homo sapiens
H296S the mutation has no effect on enzyme activity and does not inhibit tetramer assembly Homo sapiens
H324S the mutation totally prevents tetramer assembly Homo sapiens
H412S the mutation causes a complete inactivation of the enzyme with no effect on tetramer assembly or binding of the tetramer to poly(L-proline), role in the binding of Fe2+ to a catalytic site Homo sapiens
H483S the mutation causes a complete inactivation of the enzyme with no effect on tetramer assembly or binding of the tetramer to poly(L-proline), role in the binding of Fe2+ to a catalytic site Homo sapiens
H501S the mutation reduces the enzyme activity to about 4% with no effect on tetramer assembly or binding of the tetramer to poly(L-proline), role in the binding of Fe2+ to a catalytic site, the Km values for Fe2+, ascorbate and the peptide substrate with the mutant are identical to those with the wild-type enzyme, but the Km for 2-oxoglutarate is about 2.5fold higher. The main difference is that the Vmax determined from kinetic plots is consistently less than about 5% of that of the wild-type enzyme Homo sapiens
H63S the mutation has no effect on enzyme activity and does not inhibit tetramer assembly Homo sapiens
N96Q/N242Q the amount of enzyme activity observed with the double mutant alpha subunit is identical to that of the wild-type enzyme, the size of the double mutant alpha subunit is distinctly smaller than that of either the diglycosylated or monoglycosylated alpha subunit present in the wild-type enzyme, the difference being consistent with loss of all the carbohydrate Homo sapiens

Inhibitors

Inhibitors Comment Organism Structure
Pyridine 2,4-dicarboxylate inhibits wild-type enzyme and enzyme tetramer containing the histidine 501 to serine mutant alpha subunit Homo sapiens

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.018
-
(Pro-Pro-Gly)10 wild-type enzyme tetramer and enzyme tetramer containing the H501S mutant alpha subunit Homo sapiens
0.05
-
2-oxoglutarate wild-type enzyme tetramer Homo sapiens
0.15
-
2-oxoglutarate enzyme tetramer containing the H501S mutant alpha subunit Homo sapiens

Metals/Ions

Metals/Ions Comment Organism Structure
Fe2+ enzyme tetramer containing the histidine 501 to serine mutant alpha subunit: Km 0.005 mM Homo sapiens

Organism

Organism UniProt Comment Textmining
Homo sapiens
-
-
-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
enzyme activity of Triton X-100 extracts from cells expressing various mutant alpha subunits together with the wild-type protein disulfide-isomerase/beta subunit Homo sapiens

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(Pro-Pro-Gly)10 + 2-oxoglutarate + O2
-
Homo sapiens ? + succinate + CO2
-
?
(Pro-Pro-Gly)n + 2-oxoglutarate + O2 n: 1,5,10 Homo sapiens (Pro-4-hydroxy-Pro-Gly)n + succinate + CO2 n: 1,5,10 ?

Cofactor

Cofactor Comment Organism Structure
ascorbate Km for wild-type enzyme tetramer: 0.33 mM, enzyme tetramer containing the histidine 501 to serine mutant alpha subunit, Km: 0.4 mM Homo sapiens

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.002
-
Pyridine 2,4-dicarboxylate wild-type enzyme tetramer Homo sapiens
0.005
-
Pyridine 2,4-dicarboxylate enzyme tetramer containing the histidine 501 to serine mutant alpha subunit Homo sapiens