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Reaction specificity in pyridoxal phosphate enzymes

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Abstract

Pyridoxal phosphate enzymes catalyze a wide variety of reaction types on amines and amino acids, generally by stabilizing carbanionic intermediates. This makes them very useful in cellular metabolism, but it also creates problems in controlling the reaction pathway that a given enzyme follows, i.e., in controlling reaction specificity. Stereoelectronic effects have been proposed to play a major role in determining the bond to Cα that gets broken in the external aldimine intermediate that is common to all PLP enzymes. Here, we discuss our work on dialkylglycine decarboxylase aimed at providing direct evidence for stereoelectronic control of external aldimine reactivity. Once a bond to Cα has been broken to form the carbanionic intermediate, enzymes must also carefully control the fate of this reactive species. Our studies with alanine racemase suggest that the enzyme selectively destabilizes the carbanionic quinonoid intermediate to promote higher racemization specificity by avoiding transamination side reactions.

Section snippets

Model studies

The degree to which the Schiff base vs. the pyridine ring contributes to the stabilization of Cα carbanions has been debated recently. Computational studies performed by our group [5] and that of Bach [6] both predict that the pyridine ring does not play the largest role in carbanion stabilization as is commonly accepted in the literature [1], [7], [8]. Our studies used semi-empirical molecular orbital methods (PM3) to study both model aldimines and the intermediates formed in the active site

Dialkylglycine decarboxylase

This unusual and interesting PLP enzyme catalyzes the two half-reactions shown in Fig. 5A. The first is a decarboxylation reaction where 2,2-dialkylglycines lose CO2 to give a carbanion that is subsequently protonated on C4′ of the coenzyme instead of the proton replacing the CO2 on Cα of the amino acid substrate, as typical decarboxylases do. This leads to the ketone product and the PMP form of the coenzyme, as occurs in aminotransferase mechanisms. The second half-reaction is a classical

Alanine racemase

The racemization of alanine is important to bacterial survival since d-alanine is a component of the peptidoglycan layer of the cell wall structure. The X-ray structure [25], [26], [27] shows several interesting active site features. Among these are the disposition of Lys39 (which forms the Schiff base with PLP in the resting enzyme) and Tyr265 on opposite faces of the substrate Cα. This is readily seen in the structure of the complex of alanine phosphonate with alanine racemase [27], which is

References (36)

  • M.D. Toney

    Computational studies on nonenzymatic and enzymatic pyridoxal phosphate catalyzed decarboxylations of 2-aminoisobutyrate

    Biochemistry

    (2001)
  • R.D. Bach et al.

    Influence of electrostatic effects on activation barriers in enzymatic reactions: Pyridoxal 5′-phosphate-dependent decarboxylation of alpha-amino acids

    J. Am. Chem. Soc.

    (1999)
  • A.C. Eliot et al.

    Pyridoxal phosphate enzymes: mechanistic, structural, and evolutionary considerations

    Annu. Rev. Biochem.

    (2004)
  • P.D. van Poelje et al.

    Pyruvoyl-dependent enzymes

    Annu. Rev. Biochem.

    (1990)
  • C.Y. Lai et al.

    Aldolase: a model for enzyme structure–function relationships

    Essays Biochem.

    (1972)
  • D.S. Auld et al.

    J. Am. Chem. Soc.

    (1967)
  • R.F. Zabinski et al.

    Metal ion inhibition of nonenzymatic pyridoxal phosphate catalyzed decarboxylation and transamination

    J. Am. Chem. Soc.

    (2001)
  • J.E. Dixon et al.

    Comparison of the rate constants for general base catalyzed prototropy and racemization of the aldimine species formed from 3-hydroxypyridine-4-carboxaldehyde and alanine

    Biochemistry

    (1973)
  • Cited by (0)

    This work was supported by Grant GM54779 from the National Institutes of Health.

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