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Literature summary for 1.21.1.1 extracted from

  • Buss, J.M.; McTamney, P.M.; Rokita, S.E.
    Expression of a soluble form of iodotyrosine deiodinase for active site characterization by engineering the native membrane protein from Mus musculus (2012), Protein Sci., 21, 351-361.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 1.21.1.1

Application

Application Comment Organism
synthesis high expression of a truncated derivative lacking the membrane domain, residues 2-33, at its N-terminal is observed in Sf9 cells, whereas expression in Pichia pastoris remains low despite codon optimization. The desired expression in Escherichia coli can be achieved after replacing the two conserved Cys residues of the deiodinase with Ala and fusing the resulting protein to thioredoxin. This construct provides abundant enzyme for crystallography and mutagenesis Mus musculus

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Sf9 insect cells and in Escherichia coli Mus musculus

Crystallization (Commentary)

Crystallization (Comment) Organism
hanging dop vapor diffusion method, using 0.2 M ammonium acetate, 0.1 M BisTris (pH 6.5), and 45% v/v 2-methyl-2,4-pentanediol Mus musculus
structure of a truncated derivative lacking the membrane domain, residues 2-33, at its N-terminal, and its complex with substrate monoiodotyrosine. In the absence of substrate, the active site appears very accessible to solvent due to a lack of detectable structure in two surrounding regions of the polypeptide. In the presence of substrate, an active site lid comprised of a helix and loop is detected from the diffraction data. This lid effectively sequesters the substrate-flavin complex from solvent Mus musculus

Protein Variants

Protein Variants Comment Organism
E153Q the mutant exhibits no measurable binding affinity for 3-chloro-L-tyrosine Mus musculus
E153Q mutation reduces the deiodinase activity to an undetectable level. Mutant exhibits no measurable binding affinity for the substrate Mus musculus
K178Q upon expression in Escherichia coli, inactive and insoluble Mus musculus
Y157F the mutation weakens the binding of 3-chloro-L-tyrosine by 20fold Mus musculus
Y157F lack of the phenolic -OH of Y157F increases the kcat and KM values for deiodination by more than sevenfold and decreases the kcat/KM value more modestly by less than 40% Mus musculus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.019
-
 3,5-diiodo-L-tyrosine enzyme expressed in Pichia pastoris, pH not specified in the publication, temperature not specified in the publication Mus musculus
0.04
-
 3,5-diiodo-L-tyrosine enzyme expressed in Escherichia coli and lacking the transmembrane domain, pH not specified in the publication, temperature not specified in the publication Mus musculus
0.44
-
 3,5-diiodo-L-tyrosine mutant Y157F, expressed in Escherichia coli and lacking the transmembrane domain, pH not specified in the publication, temperature not specified in the publication Mus musculus

Localization

Localization Comment Organism GeneOntology No. Textmining
soluble
-
 Mus musculus
-
-

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
30000
-
 x * 30000, SDS-PAGE Mus musculus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
3,5-diiodo-L-tyrosine + NADPH + H+ Mus musculus
-
 3-iodo-L-tyrosine + NADP+ + I-
-
 ?
3-iodo-L-tyrosine + NADPH + H+ Mus musculus
-
 L-tyrosine + NADP+ + I-
-
 ?

Organism

Organism UniProt Comment Textmining
Mus musculus Q9DCX8
-
-
Mus musculus Q9DCX8 isoform iodotyrosine dehalogenase 1
-

Purification (Commentary)

Purification (Comment) Organism
HiTrap column chromatography Mus musculus

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3,5-diiodo-L-tyrosine + NADPH + H+
-
 Mus musculus 3-iodo-L-tyrosine + NADP+ + I-
-
 ?
3-iodo-L-tyrosine + NADPH + H+
-
 Mus musculus L-tyrosine + NADP+ + I-
-
 ?

Subunits

Subunits Comment Organism
? x * 30000, SDS-PAGE Mus musculus

Synonyms

Synonyms Comment Organism
iodotyrosine dehalogenase 1
-
 Mus musculus
iodotyrosine deiodinase
-
 Mus musculus
IYD
-
 Mus musculus
lyd
-
 Mus musculus

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.12
-
 3,5-diiodo-L-tyrosine enzyme expressed in Pichia pastoris, pH not specified in the publication, temperature not specified in the publication Mus musculus
0.16
-
 3,5-diiodo-L-tyrosine enzyme expressed in Escherichia coli and lacking the transmembrane domain, pH not specified in the publication, temperature not specified in the publication Mus musculus
1.08
-
 3,5-diiodo-L-tyrosine mutant Y157F, expressed in Escherichia coli and lacking the transmembrane domain, pH not specified in the publication, temperature not specified in the publication Mus musculus

Cofactor

Cofactor Comment Organism Structure
FMN
-
 Mus musculus
FMNH2
-
 Mus musculus

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.5
-
 3,5-diiodo-L-tyrosine mutant Y157F, expressed in Escherichia coli and lacking the transmembrane domain, pH not specified in the publication, temperature not specified in the publication Mus musculus
3.83
-
 3,5-diiodo-L-tyrosine enzyme expressed in Escherichia coli and lacking the transmembrane domain, pH not specified in the publication, temperature not specified in the publication Mus musculus
6
-
 3,5-diiodo-L-tyrosine enzyme expressed in Pichia pastoris, pH not specified in the publication, temperature not specified in the publication Mus musculus