Preparation of the lactate oxidase apoenzyme and studies on the binding of flavin mononucleotide to the apoenzyme

Choong, Y.S.; Shepherd, Maxwell G.; Sullivan, Patrick A.

doi:10.1042/bj1450037

Cited by 44 publications

(26 citation statements)

References 17 publications

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“…Another conventional procedure of apoprotein preparation is to partially unfold the holoprotein with guanidinium hydrochloride [121,122] or urea [21,123,124]. A disadvantage of this method is that one needs to find conditions in which the partially unfolded apoprotein is capable of refolding.…”

Section: Conventional Methodsmentioning

confidence: 99%

Deflavination and reconstitution of flavoproteins

Hefti¹,

Vervoort²,

Berkel³

2003

European Journal of Biochemistry

119

104

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Flavoproteins are ubiquitous redox proteins that are involved in many biological processes. In the majority of flavoproteins, the flavin cofactor is tightly but noncovalently bound. Reversible dissociation of flavoproteins into apoprotein and flavin prosthetic group yields valuable insights in flavoprotein folding, function and mechanism. Replacement of the natural cofactor with artificial flavins has proved to be especially useful for the determination of the solvent accessibility, polarity, reaction stereochemistry and dynamic behaviour of flavoprotein active sites. In this review we summarize the advances made in the field of flavoprotein deflavination and reconstitution. Several sophisticated chromatographic procedures to either deflavinate or reconstitute the flavoprotein on a large scale are discussed. In a subset of flavoproteins, the flavin cofactor is covalently attached to the polypeptide chain. Studies from riboflavindeficient expression systems and site-directed mutagenesis suggest that the flavinylation reaction is a post-translational, rather than a cotranslational, process. These genetic approaches have also provided insight into the mechanism of covalent flavinylation and the rationale for this atypical protein modification.

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Section: Conventional Methodsmentioning

confidence: 99%

Deflavination and reconstitution of flavoproteins

Hefti¹,

Vervoort²,

Berkel³

2003

European Journal of Biochemistry

119

104

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“…Thus, while with bacterial lactate oxidase [30,34] and with the yeast enzyme [32] the redox coenzyme is bound tightly, with the glycollate oxidases of plant origin coenzyme dissociation and comparatively weak binding appear to predominate.…”

Section: Inactivation Of the Enzyme With A-hydroxybutynoic Acidmentioning

confidence: 98%

Studies on glycollate oxidase from pea leaves determination of stereospecificity and mode of inhibition by α-hydroxybutynoate

Fendrich¹,

Ghisla²

1982

Biochimica et Biophysica Acta (BBA) - Protein Structure and Mol

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“…Values obtained by sedimentation-equilibrium analysis and an approach-to-equilibrium analysis are consistent with the above analysis. Values obtained in this study by gel chromatography and previously by non-dissociating gel electrophoresis (Choong et al, 1975) are higher, but, allowing for the accuracy of these techniques, are within experimental error.…”

Section: P a Sullivan And Othersmentioning

confidence: 39%

“…Buffers, protein determinations, the enzyme assay, enzyme units and specific activity were as described previously (Choong et al, 1975).…”

Section: Methodsmentioning

confidence: 99%

“…However, we have described studies on the preparation of the apoenzyme of lactate oxidase from M. smegmatis and reconstitution of this enzyme with FMN that indicated that the native enzyme is Vol. 165 composed of eight identical subunits and contains 8mol of FMN/mol (Choong et al, 1975). In the present paper we describe studies which show that lactate oxidase from M. smegmatis consists of eight protomers of mol.wt.…”

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confidence: 87%

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The structure of L-lactate oxidase from Mycobacterium smegmatis

Sullivan¹,

Soon²,

Schreurs³

et al. 1977

Biochemical Journal

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1. An improved purification was developed for L-lactate oxidase from Mycobacterium smegmatis. 2. The mol.wt. of the native enzyme by a sedimentation-equilibrium analysis was 345 000, and other ultracentrifuge methods gave values in the range 345 000-350 000. 3. An amino acid analysis, determinations of protein and flavin, a sedimentation-velocity analysis and an approach to equilibrium analysis gave values for the subunit mol.wt. in the range 43 500-47 000. 4. It was concluded that L-lactate oxidase contains eight subunits of mol.wt. 43 500. 5. Cross-linking of the subunits with dimethyl suberimidate and electron-microscopy studies were consistent with an octameric structure.

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Preparation of the lactate oxidase apoenzyme and studies on the binding of flavin mononucleotide to the apoenzyme

Cited by 44 publications

References 17 publications

Deflavination and reconstitution of flavoproteins

Deflavination and reconstitution of flavoproteins

Studies on glycollate oxidase from pea leaves determination of stereospecificity and mode of inhibition by α-hydroxybutynoate

The structure of L-lactate oxidase from Mycobacterium smegmatis

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