Cystathionine beta-lyase

 

Cystathionine beta-lyase is a member of the gamma family of PLP dependent enzymes, and cleaves C(beta)-S bonds in a variety of substrates. The enzyme is important in plant and microbial biosynthesis of methionine, catalysing the cleavage of L-cystathionine to L-homocysteine, pyruvate and ammonia.

The enzyme cleaves a carbon-sulfur bond, releasing L-homocysteine and an unstable enamine product that tautomerises to an imine form, which undergoes a hydrolytic deamination to form pyruvate and ammonia. The latter reaction can occur spontaneously. The enzyme from some sources also acts on L-cystine, forming pyruvate, ammonia and cysteine persulfide, and a number of related compounds.

 

Reference Protein and Structure

Sequence
P06721 UniProt (4.4.1.13, 4.4.1.28) IPR006233 (Sequence Homologues) (PDB Homologues)
Biological species
Escherichia coli K-12 (Bacteria) Uniprot
PDB
1cl1 - CYSTATHIONINE BETA-LYASE (CBL) FROM ESCHERICHIA COLI (1.83 Å) PDBe PDBsum 1cl1
Catalytic CATH Domains
3.40.640.10 CATHdb (see all for 1cl1)
Cofactors
Pyridoxal 5'-phosphate(2-) (1)
Click To Show Structure

Enzyme Reaction (EC:4.4.1.8)

L-cystathionine
CHEBI:17482ChEBI
+
water
CHEBI:15377ChEBI
→
ammonia
CHEBI:16134ChEBI
+
L-homocysteine
CHEBI:17588ChEBI
+
pyruvic acid
CHEBI:32816ChEBI
Alternative enzyme names: Beta-cystathionase, Cystine lyase, Cysteine lyase, Beta C-S lyase, Cystathionine L-homocysteine-lyase (deaminating), L-cystathionine L-homocysteine-lyase (deaminating),

Enzyme Mechanism

Introduction

Transaldimation of the substrate, forming a Schiff base with the PLP cofactor follows from the deprotonation of C(alpha) by Tyr111. This external aldimine has the alpha-carboxylate and sulphur atom aligned for proton transfer from the alpha-carbon to the epsilon-N of Lys210. In C-S bond cleavage, Lys210 abstracts the alpha-C proton, forming a quinoid intermediate. The residue's positive charge attracts the anionic phosphate of the coenzyme, aligning Lys210 to donate a proton to the gamma sulphur. Elimination of homocysteine generates the PLP derivative of aminoacrylate, which having accepting a proton from Tyr111 is converted to iminopropionate. This product is hydrolysed to pyruvate and ammonia outside of the enzyme.

Catalytic Residues Roles

UniProt PDB* (1cl1)
Tyr111 Tyr111A The residue acts as a base towards the substrate, leading to the formation of the external aldimine. Its pKa is modified through hydrogen bonding interactions with Arg 58. The residue is also thought to stabilise the quinoid intermediate through pi stacking interactions. Although more recent mutagenesis has suggested that teh residue may not act as a general acid/base. proton shuttle (general acid/base), electrostatic stabiliser
Lys210 (ptm) Llp210A (ptm) This lysine starts the reaction covalently bound to the PLP cofactor. It also acts as a general acid/base during the reaction. covalent catalysis, proton shuttle (general acid/base)
Asp185 Asp185A Acts to stabilise the PLP nitrogen that acts as an electron sink. electrostatic stabiliser
Arg58 Arg58A(AA) Acts to increase the nucleophilicity of Tyr111. increase nucleophilicity
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

References

  1. Clausen T et al. (1996), J Mol Biol, 262, 202-224. Crystal Structure of the Pyridoxal-5′-phosphate Dependent Cystathionine β-lyase fromEscherichia coliat 1.83 Å. DOI:10.1006/jmbi.1996.0508. PMID:8831789.
  2. Soo VW et al. (2016), J Biol Chem, 291, 19873-19887. Mechanistic and Evolutionary Insights from the Reciprocal Promiscuity of Two Pyridoxal Phosphate-dependent Enzymes. DOI:10.1074/jbc.m116.739557. PMID:27474741.
  3. Astegno A et al. (2015), Proteins, 83, 78-90. Role of active-site residues Tyr55 and Tyr114 in catalysis and substrate specificity ofCorynebacterium diphtheriaeC-S lyase. DOI:10.1002/prot.24707. PMID:25354840.
  4. Lodha PH et al. (2011), Biochemistry, 50, 9876-9885. Characterization of the Side-Chain Hydroxyl Moieties of Residues Y56, Y111, Y238, Y338, and S339 as Determinants of Specificity inE. coliCystathionine β-Lyase. DOI:10.1021/bi201090n. PMID:21958132.
  5. Lodha PH et al. (2010), Protein Sci, 19, 383-391. Characterization of site-directed mutants of residues R58, R59, D116, W340 and R372 in the active site ofE. colicystathionine β-lyase. DOI:10.1002/pro.308. PMID:20014435.
  6. Ejim LJ et al. (2007), J Med Chem, 50, 755-764. Inhibitors of Bacterial Cystathionine β-Lyase:  Leads for New Antimicrobial Agents and Probes of Enzyme Structure and Function. DOI:10.1021/jm061132r. PMID:17300162.
  7. Messerschmidt A et al. (2003), Biol Chem, 384, 373-386. Determinants of Enzymatic Specificity in the Cys-Met-Metabolism PLP-Dependent Enzyme Family: Crystal Structure of Cystathionine γ-Lyase from Yeast and Intrafamiliar Structure Comparison. DOI:10.1515/bc.2003.043. PMID:12715888.
  8. Clausen T et al. (1997), Biochemistry, 36, 12633-12643. Slow-Binding Inhibition ofEscherichia coliCystathionine β-Lyase byl-Aminoethoxyvinylglycine:  A Kinetic and X-ray Study‡. DOI:10.1021/bi970630m. PMID:9376370.

Step 1.

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Catalytic Residues Roles

Residue Roles
Tyr111A electrostatic stabiliser, proton shuttle (general acid/base)
Llp210A (ptm) covalent catalysis, proton shuttle (general acid/base)
Asp185A electrostatic stabiliser
Arg58A(AA) increase nucleophilicity

Chemical Components

Step 1.

Contributors

Gemma L. Holliday, Nozomi Nagano, Craig Porter