Folding pathway of the pyridoxal 5'-phosphate C-S lyase MalY from Escherichia coli.

Article Details

Citation

Copy to clipboard

Bertoldi M, Cellini B, Laurents DV, Borri Voltattorni C

Folding pathway of the pyridoxal 5'-phosphate C-S lyase MalY from Escherichia coli.

Biochem J. 2005 Aug 1;389(Pt 3):885-98.

PubMed ID

15823094 [ View in PubMed

]

Abstract

MalY from Escherichia coli is a bifunctional dimeric PLP (pyridoxal 5'-phosphate) enzyme acting as a beta-cystathionase and as a repressor of the maltose system. The spectroscopic and molecular properties of the holoenzyme, in the untreated and NaBH4-treated forms, and of the apoenzyme have been elucidated. A systematic study of the urea-induced unfolding of MalY has been monitored by gel filtration, cross-linking, ANS (8-anilino-1-naphthalenesulphonic acid) binding and by visible, near- and far-UV CD, fluorescence and NMR spectroscopies under equilibrium conditions. Unfolding proceeds in at least three stages. The first transition, occurring between 0 and 1 M urea, gives rise to a partially active dimeric species that binds PLP. The second equilibrium transition involving dimer dissociation, release of PLP and loss of lyase activity leads to the formation of a monomeric equilibrium intermediate. It is a partially unfolded molecule that retains most of the native-state secondary structure, binds significant amounts of ANS (a probe for exposed hydrophobic surfaces) and tends to self-associate. The self-associated aggregates predominate at urea concentrations of 2-4 M for holoMalY. The third step represents the complete unfolding of the enzyme. These results when compared with the urea-induced unfolding profiles of apoMalY and NaBH4-reduced holoenzyme suggest that the coenzyme group attached to the active-site lysine residue increases the stability of the dimeric enzyme. Both holo- and apo-MalY could be successfully refolded into the active enzyme with an 85% yield. Further refolding studies suggest that large misfolded soluble aggregates that cannot be refolded could be responsible for the incomplete re-activation.

DrugBank Data that Cites this Article

Drug Targets

Drug	Target	Kind	Organism	Pharmacological Action	Actions
Pyridoxal phosphate	Cystathionine gamma-lyase	Protein	Humans	Unknown	Cofactor	Details