PDBsum entry 1yjs

Transferase

PDB id: 1yjs

Contents

Protein chain

405 a.a. *

Ligands

PLP-GLY

Waters ×139

* Residue conservation analysis

PDB id:

1yjs

Name:

Transferase

Title:

K226q mutant of serine hydroxymethyltransferase from b. Stearothermophilus, complex with glycine

Structure:

Serine hydroxymethyltransferase. Chain: a. Fragment: serine methylase. Engineered: yes. Mutation: yes

Source:

Geobacillus stearothermophilus. Organism_taxid: 1422. Gene: shmt. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.

Biol. unit:

Dimer (from PDB file)

Resolution:

2.00Å R-factor: 0.203 R-free: 0.227

Authors:

S.Bhavani,V.Trivedi,V.R.Jala,H.S.Subramanya,P.Kaul,K.Purnima, V.Prakash,R.N.Appaji,H.S.Savithri

Key ref:

S.Bhavani et al. (2005). Role of Lys-226 in the catalytic mechanism of Bacillus stearothermophilus serine hydroxymethyltransferase--crystal structure and kinetic studies. Biochemistry, 44, 6929-6937. PubMed id: 15865438 DOI: 10.1021/bi047800x

Date:

15-Jan-05 Release date: 31-May-05

Protein chain

Q7SIB6  (Q7SIB6_GEOSE) -  Serine hydroxymethyltransferase from Geobacillus stearothermophilus

Seq: 419 a.a.

Struc: 405 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.2.1.2.1 - glycine hydroxymethyltransferase.
• Pathway: Folate Coenzymes
• Reaction: (6R)-5,10-methylene-5,6,7,8-tetrahydrofolate + glycine + H2O = (6S)- 5,6,7,8-tetrahydrofolate + L-serine

(6R)-5,10-methylene-5,6,7,8-tetrahydrofolate + glycine (Bound ligand (Het Group name = GLY) corresponds exactly) + H2O = (6S)- 5,6,7,8-tetrahydrofolate + L-serine

• Cofactor: Pyridoxal 5'-phosphate

Pyridoxal 5'-phosphate (Bound ligand (Het Group name = PLP) matches with 93.75% similarity)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1021/bi047800x

Biochemistry 44:6929-6937 (2005)

PubMed id: 15865438

Role of Lys-226 in the catalytic mechanism of Bacillus stearothermophilus serine hydroxymethyltransferase--crystal structure and kinetic studies.

S.Bhavani, V.Trivedi, V.R.Jala, H.S.Subramanya, P.Kaul, V.Prakash, N.Appaji Rao, H.S.Savithri.

ABSTRACT

Serine hydroxymethyltransferase (SHMT), a pyridoxal 5'-phosphate (PLP)-dependent enzyme catalyzes the reversible conversion of l-Ser and tetrahydropteroylglutamate (H(4)PteGlu) to Gly and 5,10-methylene tetrahydropteroylglutamate (CH(2)-H(4)PteGlu). Biochemical and structural studies on this enzyme have implicated several residues in the catalytic mechanism, one of them being the active site lysine, which anchors PLP. It has been proposed that this residue is crucial for product expulsion. However, in other PLP-dependent enzymes, the corresponding residue has been implicated in the proton abstraction step of catalysis. In the present investigation, Lys-226 of Bacillus stearothermophilus SHMT (bsSHMT) was mutated to Met and Gln to evaluate the role of this residue in catalysis. The mutant enzymes contained 1 mol of PLP per mol of subunit suggesting that Schiff base formation with lysine is not essential for PLP binding. The 3D structure of the mutant enzymes revealed that PLP was bound at the active site in an orientation different from that of the wild-type enzyme. In the presence of substrate, the PLP ring was in an orientation superimposable with that of the external aldimine complex of wild-type enzyme. However, the mutant enzymes were inactive, and the kinetic analysis of the different steps of catalysis revealed that there was a drastic reduction in the rate of formation of the quinonoid intermediate. Analysis of these results along with the crystal structures suggested that K-226 is responsible for flipping of PLP from one orientation to another which is crucial for H(4)PteGlu-dependent Calpha-Cbeta bond cleavage of l-Ser.