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Literature summary for 2.7.1.39 extracted from

  • Petit, C.; Kim, Y.; Lee, S.K.; Brown, J.; Larsen, E.; Ronning, D.R.; Suh, J.W.; Kang, C.M.
    Reduction of feedback inhibition in homoserine kinase (ThrB) of Corynebacterium glutamicum enhances L-threonine biosynthesis (2018), ACS omega, 3, 1178-1186 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
gene thrB, recombinant expression of His-taged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Corynebacterium glutamicum

Crystallization (Commentary)

Crystallization (Comment) Organism
purified recombinant His-tagged enzyme in apoform or with either methionine or ATPgammaS and cysteine, hanging drop vapor diffusion method, mixing of 0.002 ml of 16.2 mg/ml protein in 20 mM Tris, pH 7.5, 150 mM NaCl, 50 mM KCl, and 50 mM MgCl2 with 0.002 ml of well solution containing 0.25 M ammonium sulfate, 25% PEG 3350, and 0.1 M HEPES, pH 7.5, and equilibration against 0.1 ml of well solution, X-ray diffraction structure determination and analysis at 1.8-2.2 A resolution Corynebacterium glutamicum

Protein Variants

Protein Variants Comment Organism
A20G site-directed mutagenesis, the mutant retains wild-type enzymatic activity, with dramatically decreased feedback inhibition by L-threonine. The changes in L-threonine affinity to the CglThrB-A20G active site derive from loss of van der Waals interactions Corynebacterium glutamicum
A20L site-directed mutagenesis Corynebacterium glutamicum
A20S site-directed mutagenesis Corynebacterium glutamicum
A20V site-directed mutagenesis Corynebacterium glutamicum
additional information engineering of Corynebacterium glutamicum to increase production of L-threonine, the attempt is to render the active site of Corynebacterium glutamicum ThrB (CglThrB) more selective toward L-homoserine than L-threonine Corynebacterium glutamicum

Inhibitors

Inhibitors Comment Organism Structure
L-threonine competitive inhibition, interaction with ThrB residue A20 is important. Highly reduced inhibition of enzyme mutant A20G. The changes in L-threonine affinity to the CglThrB-A20G active site derive from loss of van der Waals interactions Corynebacterium glutamicum

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Corynebacterium glutamicum

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + L-homoserine Corynebacterium glutamicum
-
ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine Corynebacterium glutamicum LMG 3730
-
ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine Corynebacterium glutamicum BCRC 11384
-
ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine Corynebacterium glutamicum ATCC 13032
-
ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine Corynebacterium glutamicum JCM 1318
-
ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine Corynebacterium glutamicum NCIMB 10025
-
ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine Corynebacterium glutamicum DSM 20300
-
ADP + O-phospho-L-homoserine
-
?

Organism

Organism UniProt Comment Textmining
Corynebacterium glutamicum P07128
-
-
Corynebacterium glutamicum ATCC 13032 P07128
-
-
Corynebacterium glutamicum BCRC 11384 P07128
-
-
Corynebacterium glutamicum DSM 20300 P07128
-
-
Corynebacterium glutamicum JCM 1318 P07128
-
-
Corynebacterium glutamicum LMG 3730 P07128
-
-
Corynebacterium glutamicum NCIMB 10025 P07128
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-taged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and dialysis Corynebacterium glutamicum

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + L-homoserine
-
Corynebacterium glutamicum ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine
-
Corynebacterium glutamicum LMG 3730 ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine
-
Corynebacterium glutamicum BCRC 11384 ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine
-
Corynebacterium glutamicum ATCC 13032 ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine
-
Corynebacterium glutamicum JCM 1318 ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine
-
Corynebacterium glutamicum NCIMB 10025 ADP + O-phospho-L-homoserine
-
?
ATP + L-homoserine
-
Corynebacterium glutamicum DSM 20300 ADP + O-phospho-L-homoserine
-
?

Subunits

Subunits Comment Organism
More Corynebacterium glutamicum CbThrB crystal structure (PDB ID 5WAT) comparison with the crystal structure of Methanocaldococcus jannaschii ThrB with bound L-Thr or L-homoserine (MjaThrB, PDB ID 1H72 or 1H73), overview Corynebacterium glutamicum

Synonyms

Synonyms Comment Organism
CglThrB
-
Corynebacterium glutamicum
CThrB
-
Corynebacterium glutamicum
ThrB
-
Corynebacterium glutamicum

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
27
-
assay at Corynebacterium glutamicum

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.8
-
assay at Corynebacterium glutamicum

General Information

General Information Comment Organism
additional information a conserved active-site alanine residue, A20, in CThrB (CglThrB) is important for differential interactions with L-threonine and L-homoserine Corynebacterium glutamicum