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Literature summary for 1.14.11.B17 extracted from

  • Strieker, M.; Nolan, E.M.; Walsh, C.T.; Marahiel, M.A.
    Stereospecific synthesis of threo- and erythro-beta-hydroxyglutamic acid during kutzneride biosynthesis (2009), J. Am. Chem. Soc., 131, 13523-13530.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
overproduced in Escherichia coli as His7-tagged fusions Kutzneria sp. 744

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information nonhydrolyzable coenzyme A analogs are developed and used to determine the kinetic parameters for KtzO-catalyzed hydroxylation of glutamic acid bound to the carrier protein. To determine the kinetic parameters of KtzO catalyzed hydroxylation, the problem of the labile thioester bond is circumvented by using synthetic coenzyme A analogs coupled to glutamic acid where the thioester is replaced by a hydrolytically stable amide bond Kutzneria sp. 744

Metals/Ions

Metals/Ions Comment Organism Structure
Iron putative non-heme iron oxygenase, exhibit the conserved HXD/E_H iron(II)-binding motif, in the assay (NH4)2FeSO4 is used as source of the ferrous iron cofactor (0.5 mM) Kutzneria sp. 744

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
40100
-
x * 40100, SDS-PAGE Kutzneria sp. 744

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
S-(L-glutamyl)-[peptidyl-carrier protein of nonribosomal peptide synthetase KtzH] + 2-oxoglutarate + O2 Kutzneria sp. 744 the enzyme is involved in the biosynthesis of L-threo-3-hydroxy-glutamate, prior to incorporation into kutznerides (antifungal nonribosomal hexadepsipeptides) S-(threo-3-hydroxy-L-glutamyl)-[peptidyl-carrier-protein of nonribosomal peptide synthetase KtzH] + succinate + CO2
-
?

Organism

Organism UniProt Comment Textmining
Kutzneria sp. 744 A8CF77
-
-

Purification (Commentary)

Purification (Comment) Organism
-
Kutzneria sp. 744

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
additional information able to hydroxylate glutamic acid bound to a noncognate peptidyl-carrier-protein, e.g. the ninth peptidyl-carrier-protein domain of the CDA NRPS assembly line (CDA-T9). Nonhydrolyzable coenzyme A analogs are developed and used to determine the kinetic parameters for KtzO-catalyzed hydroxylation of glutamic acid bound to the carrier protein. To determine the kinetic parameters of KtzO catalyzed hydroxylation, the problem of the labile thioester bond is circumvented by using synthetic coenzyme A analogs coupled to glutamic acid where the thioester is replaced by a hydrolytically stable amide bond Kutzneria sp. 744 ?
-
?
S-(L-glutamyl)-[peptidyl-carrier protein of nonribosomal peptide synthetase KtzH] + 2-oxoglutarate + O2 the enzyme is involved in the biosynthesis of L-threo-3-hydroxy-glutamate, prior to incorporation into kutznerides (antifungal nonribosomal hexadepsipeptides) Kutzneria sp. 744 S-(threo-3-hydroxy-L-glutamyl)-[peptidyl-carrier-protein of nonribosomal peptide synthetase KtzH] + succinate + CO2
-
?
S-(L-glutamyl)-[peptidyl-carrier protein of nonribosomal peptide synthetase KtzH] + 2-oxoglutarate + O2 D-Glu, L-Glu and [peptidyl-carrier-protein]-D-glutamyl thioester are not accepted as substrates. Proposed reaction mechanism: the stand-alone adenylation domain KtzN first activates L-glutamic acid as amino acyl adenylate, which then can be transferred site-specifically to the third peptidyl-carrier-protein domain of KtzH. The specificity of this interaction is mediated by the presence of the truncated A domain (A*) as well as hydroxylase KtzO. Both A* and a hydroxylase KtzO are required for Glu-AMP transfer. Subsequently, the PCP-bound glutamic acid is hydroxylated by KtzO to afford L-threo-3-hydroxyglutamic acid Kutzneria sp. 744 S-(threo-3-hydroxy-L-glutamyl)-[peptidyl-carrier-protein of nonribosomal peptide synthetase KtzH] + succinate + CO2
-
?

Subunits

Subunits Comment Organism
? x * 40100, SDS-PAGE Kutzneria sp. 744

Synonyms

Synonyms Comment Organism
KtzO
-
Kutzneria sp. 744

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
28
-
assay at Kutzneria sp. 744

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
additional information
-
additional information nonhydrolyzable coenzyme A analogs are developed and used to determine the kinetic parameters for KtzO-catalyzed hydroxylation of glutamic acid bound to the carrier protein. To determine the kinetic parameters of KtzO catalyzed hydroxylation, the problem of the labile thioester bond is circumvented by using synthetic coenzyme A analogs coupled to glutamic acid where the thioester is replaced by a hydrolytically stable amide bond Kutzneria sp. 744

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7
-
assay at Kutzneria sp. 744

General Information

General Information Comment Organism
physiological function the enzyme is involved in the biosynthesis of L-threo-3-hydroxy-glutamate, prior to incorporation into kutznerides (antifungal nonribosomal hexadepsipeptides) Kutzneria sp. 744

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
additional information
-
additional information nonhydrolyzable coenzyme A analogs are developed and used to determine the kinetic parameters for KtzO-catalyzed hydroxylation of glutamic acid bound to the carrier protein. To determine the kinetic parameters of KtzO catalyzed hydroxylation, the problem of the labile thioester bond is circumvented by using synthetic coenzyme A analogs coupled to glutamic acid where the thioester is replaced by a hydrolytically stable amide bond Kutzneria sp. 744