PDBsum entry 2xbn

Transferase

PDB id: 2xbn

Contents

Protein chain

398 a.a. *

Ligands

PMP

Metals

_MG

Waters ×298

* Residue conservation analysis

PDB id:

2xbn

Name:

Transferase

Title:

Inhibition of the plp-dependent enzyme serine palmitoyltransferase by cycloserine: evidence for a novel decarboxylative mechanism of inactivation

Structure:

Serine palmitoyltransferase. Chain: a. Fragment: residues 2-420. Synonym: spt, spt1. Engineered: yes

Source:

Sphingomonas paucimobilis. Organism_taxid: 13689. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

1.40Å R-factor: 0.157 R-free: 0.181

Authors:

J.Lowther,B.A.Yard,K.A.Johnson,L.G.Carter,V.T.Bhat,M.C.C.Raman, D.J.Clarke,B.Ramakers,S.A.Mcmahon,J.H.Naismith,D.J.Campopiano

Key ref:

J.Lowther et al. (2010). Inhibition of the PLP-dependent enzyme serine palmitoyltransferase by cycloserine: evidence for a novel decarboxylative mechanism of inactivation. Mol Biosyst, 6, 1682-1693. PubMed id: 20445930

Date:

13-Apr-10 Release date: 19-May-10

Protein chain

Q93UV0  (SPT_SPHPI) -  Serine palmitoyltransferase from Sphingomonas paucimobilis

Seq: 420 a.a.

Struc: 398 a.a.

Enzyme reactions

• Enzyme class: E.C.2.3.1.50 - serine C-palmitoyltransferase.
• Reaction: L-serine + hexadecanoyl-CoA + H⁺ = 3-oxosphinganine + CO2 + CoA
• Cofactor: Pyridoxal 5'-phosphate

Pyridoxal 5'-phosphate (Bound ligand (Het Group name = PMP) matches with 88.24% similarity)

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

reference

Mol Biosyst 6:1682-1693 (2010)

PubMed id: 20445930

Inhibition of the PLP-dependent enzyme serine palmitoyltransferase by cycloserine: evidence for a novel decarboxylative mechanism of inactivation.

J.Lowther, B.A.Yard, K.A.Johnson, L.G.Carter, V.T.Bhat, M.C.Raman, D.J.Clarke, B.Ramakers, S.A.McMahon, J.H.Naismith, D.J.Campopiano.

ABSTRACT

Cycloserine (CS, 4-amino-3-isoxazolidone) is a cyclic amino acid mimic that is known to inhibit many essential pyridoxal 5'-phosphate (PLP)-dependent enzymes. Two CS enantiomers are known; d-cycloserine (DCS, also known as Seromycin) is a natural product that is used to treat resistant Mycobacterium tuberculosis infections as well as neurological disorders since it is a potent NMDA receptor agonist, and l-cycloserine (LCS) is a synthetic enantiomer whose usefulness as a drug has been hampered by its inherent toxicity arising through inhibition of sphingolipid metabolism. Previous studies on various PLP-dependent enzymes revealed a common mechanism of inhibition by both enantiomers of CS; the PLP cofactor is disabled by forming a stable 3-hydroxyisoxazole/pyridoxamine 5'-phosphate (PMP) adduct at the active site where the cycloserine ring remains intact. Here we describe a novel mechanism of CS inactivation of the PLP-dependent enzyme serine palmitoyltransferase (SPT) from Sphingomonas paucimobilis. SPT catalyses the condensation of l-serine and palmitoyl-CoA, the first step in the de novo sphingolipid biosynthetic pathway. We have used a range of kinetic, spectroscopic and structural techniques to postulate that both LCS and DCS inactivate SPT by transamination to form a free pyridoxamine 5'-phosphate (PMP) and beta-aminooxyacetaldehyde that remain bound at the active site. We suggest this occurs by ring opening of the cycloserine ring followed by decarboxylation. Enzyme kinetics show that inhibition is reversed by incubation with excess PLP and that LCS is a more effective SPT inhibitor than DCS. UV-visible spectroscopic data, combined with site-directed mutagenesis, suggest that a mobile Arg(378) residue is involved in cycloserine inactivation of SPT.