1. Analogs of 1-phosphonooxy-2,2-dihydroxy-3-oxo-5-(methylthio)pentane, an acyclic intermediate in the methionine salvage pathway: a new preparation and characterization of activity with E1 enolase/phosphatase from Klebsiella oxytoca.
- Author
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Zhang Y, Heinsen MH, Kostic M, Pagani GM, Riera TV, Perovic I, Hedstrom L, Snider BB, and Pochapsky TC
- Subjects
- Amino Acid Sequence, Base Sequence, DNA Primers, Magnetic Resonance Spectroscopy, Methionine chemistry, Models, Molecular, Molecular Sequence Data, Phosphoric Monoester Hydrolases chemistry, Sequence Homology, Amino Acid, Spectrometry, Mass, Fast Atom Bombardment, Spectrophotometry, Ultraviolet, Klebsiella oxytoca enzymology, Methionine metabolism, Organophosphates chemistry, Pentanes chemistry, Phosphopyruvate Hydratase metabolism, Phosphoric Monoester Hydrolases metabolism
- Abstract
The methionine salvage pathway allows the in vivo recovery of the methylthio moiety of methionine upon the formation of methylthioadenosine (MTA) from S-adenosylmethionine (SAM). The Fe(II)-containing form of acireductone dioxygenase (ARD) catalyzes the penultimate step in the pathway in Klebsiella oxytoca, the oxidative cleavage of the acireductone 1,2-dihydroxy-3-oxo-5-(methylthio)pent-1-ene (2) by dioxygen to give formate and 2-oxo-4-(methylthio)butyrate (3). The Ni(II)-bound form (Ni-ARD) catalyzes an off-pathway shunt, forming 3-(methylthio)propionate (4), carbon monoxide, and formate. Acireductone 2 is formed by the action of another enzyme, E1 enolase/phosphatase, on precursor 1-phosphonooxy-2,2-dihydroxy-3-oxo-5-methylthiopentane (1). Simple syntheses of several analogs of 1 are described, and their activity as substrates for E1 enolase/phosphatase characterized. A new bacterial overexpression system and purification procedure for E1, a member of the haloacid dehalogenase (HAD) superfamily, is described, and further characterization of the enzyme presented.
- Published
- 2004
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