Purification and characterization of l-allo-threonine aldolase from Aeromonas jandaei DK-39

Abstract

L-allo-Threonine aldolase (L-allo-threonine acetaldehyde-lyase), which exhibited specificity for L-allo-threonine but not for L-threonine, was purified from a cell-free extract of Aeromonas jandaei DK-39. The purified enzyme catalyzed the aldol cleavage reaction of L-allo-threonine (K(m) = 1.45 mM, Vmax = 45.2 mumol min-1 mg-1). The activity of the enzyme was inhibited by carbonyl reagents, which suggests that pyridoxal-5'-phosphate participates in the enzymatic reaction. The enzyme does not act on either L-serine or L-threonine, and thus it can be distinguished from serine hydroxy-methyltransferase (L-serine:tetrahydrofolate 5,10-hydroxy-methyltransferase, EC 2.1.2.1) or L-threonine aldolase (EC 4.1.2.5).

Expanding the threonine aldolase toolbox for the asymmetric synthesis of tertiary α-amino acids

“…Interestingly, the kinetic parameters for the cleavage of non-natural α-methyl-β-phenylserine lie in a similar range or are even higher as for the cleavage of natural L-threonine in the case of Lspecific TAs. This can be explained by the high stereospecificity of these enzymes toward the L-allo-threonine isomer compared to L-threonine, as was shown before (Kataoka et al 1997). In contrast, the catalytic efficiency k cat /K m for the D-specific enzymes was lower for the retro-aldol cleavage of α-methyl-β-phenylserine (e.g., k cat /K m 0.19 min −1 mM −1 for DTA_Pa) compared to the reaction with natural substrate D-threonine (k cat /K m 1.91 min − 1 mM − 1 for DTA_Pa) and especially to the cleavage of D-β-phenylserine (k cat /K m 47.2 min −1 mM −1 for DTA_Pa).…”

Großes Kino

“…Recently a biocatalytic approach to enantiopure β‐hydroxy‐α‐amino acids has been developed by application of threonine aldolases and investigated in detail in our group 8. In a search for potential catalysts for the asymmetric synthesis of α,α‐dialkyl‐α‐amino acids, we found two natural threonine aldolases that catalyze the cleavage of racemic α‐methylthreonine ( 1 ) to produce acetaldehyde and D ‐alanine: an L ‐ allo‐ threonine aldolase from Aeromonas jandaei ( L ‐TA)9 and a D ‐threonine aldolase from Pseudomonas sp. ( D ‐TA; Scheme ).…”

Synthesis of 4-Hydroxyisoleucine by the Aldolase–Transaminase Coupling Reaction and Basic Characterization of the Aldolase fromArthrobacter simplexAKU 626

“…There have been some reports on PLPdependent aldolases. [13][14][15][16] It is not clear that the Arthrobacter aldolase requires PLP as a coenzyme, because the transaminase in the reaction mixture also requires PLP as a cofactor. Further characterization, cloning, and expression of the aldolase are in progress.…”