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Literature summary for 1.1.1.27 extracted from

  • Cook, W.J.; Senkovich, O.; Hernandez, A.; Speed, H.; Chattopadhyay, D.
    Biochemical and structural characterization of Cryptosporidium parvum lactate dehydrogenase (2015), Int. J. Biol. Macromol., 74, 608-619.
    View publication on PubMed

Application

Application Comment Organism
analysis CpLDH with APAD+ may be useful as a diagnostic tool for detection of protozoan parasite Cryptosporidium Cryptosporidium parvum

Cloned(Commentary)

Cloned (Comment) Organism
gene LDH1, recombinant expression of the enzyme in Escherichia coli strain Rosetta (DE3)pLysS, molecular replacement and modelling using the structure of PfLDH, PDB ID 1T2D Cryptosporidium parvum

Crystallization (Commentary)

Crystallization (Comment) Organism
purified recombinant enzyme as apo-enzyme and in four ternary complexes, The ternary complexes capture the enzyme bound to NAD/NADH or its 3-acetylpyridine analogue in the cofactor binding pocket, while the substrate binding site is occupied by one of the following ligands: lactate, pyruvate or oxamate. X-ray diffraction structure determination and analysis at 2.0-2.3 A resolution Cryptosporidium parvum

Inhibitors

Inhibitors Comment Organism Structure
gossypol
-
Cryptosporidium parvum
additional information CpLDH does not display any measurable inhibition to pyruvate concentrations up to at least 20 mM in contrast to many other LDHs Cryptosporidium parvum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetics Cryptosporidium parvum
0.0018
-
(S)-lactate pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.0085
-
APAD+ pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.0166
-
APADH pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.0169
-
NADH pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.0303
-
NAD+ pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.1973
-
pyruvate pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
(S)-lactate + NAD+ Cryptosporidium parvum
-
pyruvate + NADH + H+
-
r
pyruvate + NADH + H+ Cryptosporidium parvum
-
(S)-lactate + NAD+
-
r

Organism

Organism UniProt Comment Textmining
Cryptosporidium parvum Q9GT92
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant enzyme from Escherichia coli strain Rosetta (DE3)pLysS by ammonium sulfate fractionation, gel filtration, anion exchange chromatography, and ultrafiltration Cryptosporidium parvum

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(S)-lactate + APAD+
-
Cryptosporidium parvum pyruvate + APADH + H+
-
r
(S)-lactate + NAD+
-
Cryptosporidium parvum pyruvate + NADH + H+
-
r
pyruvate + APADH + H+
-
Cryptosporidium parvum (S)-lactate + APAD+
-
r
pyruvate + NADH + H+
-
Cryptosporidium parvum (S)-lactate + NAD+
-
r

Subunits

Subunits Comment Organism
More the ternary complexes capture the enzyme bound to NAD/NADH or its 3-acetylpyridine analog in the cofactor binding pocket, while the substrate binding site is occupied by one of the following ligands: lactate, pyruvate or oxamate Cryptosporidium parvum

Synonyms

Synonyms Comment Organism
lactate dehydrogenase
-
Cryptosporidium parvum
LDH1
-
Cryptosporidium parvum

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Cryptosporidium parvum

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.000045
-
(S)-lactate pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.000055
-
NAD+ pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.000248
-
APAD+ pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.00208
-
NADH pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.00209
-
pyruvate pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.00386
-
APADH pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
5 5.5 reduction of pyruvate Cryptosporidium parvum
8.5 9.5 oxidation of lactate Cryptosporidium parvum

pH Range

pH Minimum pH Maximum Comment Organism
4 8 over 40% of maximal activity for reduction of pyruvate within this range Cryptosporidium parvum
8 9.5 20% of maximal activity for oxidation of lactate at pH 8.0, 90% at pH 8.5, and maximal activity at pH 8.5-9.5, inactive at pH 7.5 Cryptosporidium parvum

Cofactor

Cofactor Comment Organism Structure
3-acetyl-pyridine adenine dinucleotide i.e. APAD+, 3-acetylpyridine adenine dinucleotide, analogue of NAD+ Cryptosporidium parvum
additional information Cryptosporidium parvum lactate dehydrogenase prefers the acetylpyridine analogue of NADH as a cofactor. NADH/APADH and substrate binding site structures, overview. replacement of the conserved serine residue by Met163 in CpLDH allows a better packing environment for the acetyl of APAD+ Cryptosporidium parvum
NAD+
-
Cryptosporidium parvum
NADH
-
Cryptosporidium parvum

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.0116
-
gossypol pH 5.5, 25°C, recombinant enzyme, with NADH and pyruvate Cryptosporidium parvum

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.000033
-
(S)-lactate pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.00183
-
NAD+ pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.0105
-
pyruvate pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.0292
-
APAD+ pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.1235
-
NADH pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum
0.2326
-
APADH pH 5.5, 25°C, recombinant enzyme Cryptosporidium parvum