O-succinyl-L-homoserine succinate-lyase (adding cysteine)
This enzyme was transferred from E.C. 4.2.99.9 to E.C. 2.5.1.48. This enzyme belongs to the cystathionine beta-lyase subclass of the type-I PLP-dependent enzyme superfamily (Aminotransferase superfamily) (see [5] & [9]). This enzyme catalyzes the following reactions: (A) Formation of external aldimine (with amine group of O-succinyl-L-homocysteine), (B) Isomerization (change in the position of double-bond), (C) Elimination of carboxyl ester group, forming double-bonded carbon atoms, leading to beta-gamma unsaturated intermediate, (D) Addition of the second substrate, sulfide, to the double-bonded beta-carbon of the unsaturated intermediate, (E) Isomerization (change in the position of double-bond), (F) Formation of internal aldimine, leading to the elimination of the product from PLP. According to the literature [4], [7] & [10], these reactions proceed as follows: Here, Tyr108 (of 1qgn) from the adjacent chain modulates the function of Lys261 as base-acid during catalysis. Moreover, Asp236 stabilizes the positively charged pyridine nitrogen of PLP, increasing its electrophilic character (see [4]). (A) Formation of external aldimine (with amine group of O-succinyl-L-homocysteine): (A1) Lys261 (of 1qgn) is covalently bound to the PLP cofactor. (Here, Arg110 (from the adjacent chain) and the protonated aldimine of the PLP lower the pKa of the sidechain of Tyr163) (A2) Tyr163 acts as a general base, which deprotonates the amine group of the first substrate, O-succinyl-L-homocysteine. (A3) The deprotonated amine group makes a nucleophilic attack on the C4' atom of the PLP, which is bound to Lys261, leading to a tetrahedral intermediate. (A4) A proton on the amine from the first substrate must be transferred to the leaving nitrogen atom, somehow. (A5) A lone pair on the amine group makes a nucleophilic attack on the C4' atom of the PLP, leading to the formation of the double-bond and the release of the leaving group. (B) Isomerization (change in the position of double-bond) [This reaction does not involve quinoid intermediate. see [4]]: (B1) Lys261 acts as a general base, to deprotonate the alpha-carbon of the first substrate, now covalently bound to the PLP cofactor, forming a carbanionic inermediate. (B2) Lys261 acts as a general acid, to protonate the C4' atom of the PLP, leading to the formation of PLP-substrate ketimine. (C) Elimination of carboxyl ester group, forming double-bonded carbon atoms, leading to beta-gamma unsaturated intermediate: (C1) Lys261 acts as a general base, to deprotonate the beta-carbon (deprotonation site) of the ketimine intermediate, leading to the formation of transition state, alpha-beta unsaturated ketimine. (C2) Tyr163 acts as a general acid, to protonate the ester bond of the eliminated group, succinate, resulting in the formation of a beta-gamma unsaturated intermediate and the release of succinate. (D) Addition of the second substrate, sulfide, to the double-bonded beta-carbon of the beta-gamma unsaturated intermediate: (D1) The sulfur atom of the second substrate, cysteine, makes a nucleophilic attack on the gamma-carbon (sp2) of the intermediate. (D2) Lys261 acts as a general acid, to protonate the beta-carbon of the intermdiate, resulting in the formation of the PLP product ketimine. (E) Isomerization (change in the position of double-bond): (E1) Lys261 acts as a general base, to deprotonate the C4' atom of the PLP, forming a carbanionic intermediate. (E2) Lys261 acts as a general acid, to protonate the alpha-carbon of the carbanionic intermediate, returning to the external aldimine. (F) Formation of internal aldimine, leading to the elimination of the product from PLP: Although this final reaction has not been elucidated in the literature, Lys261 acts as a nucleophile to form the internal aldimine.
Proton retention in the gamma-elimination reaction catalysed by cystathionine gamma-synthase.
Crystal structures of cystathionine gamma-synthase inhibitor complexes rationalize the increased affinity of a novel inhibitor.
Homocysteine biosynthesis in green plants. O-Phosphorylhomoserine as the physiological substrate for cystathionine gamma-synthase.
Cloning, purification, crystallization, and preliminary X-ray diffraction analysis of cystathionine gamma-synthase from E. coli.
Crystal structure of Escherichia coli cystathionine gamma-synthase at 1.5 A resolution.
Cloning, purification and characterisation of cystathionine gamma-synthase from Nicotiana tabacum.
The crystal structure of cystathionine gamma-synthase from Nicotiana tabacum reveals its substrate and reaction specificity.
The manifold of vitamin B6 dependent enzymes.
Identification of a short highly conserved amino acid sequence as the functional region required for posttranscriptional autoregulation of the cystathionine gamma-synthase gene in Arabidopsis.
Structure, evolution and action of vitamin B6-dependent enzymes.
Mechanisms to account for maintenance of the soluble methionine pool in transgenic Arabidopsis plants expressing antisense cystathionine gamma-synthase cDNA.