Purification and characterization of pyruvate kinase from schizosaccharomyces pombe: evidence for an unusual quaternary structure

Nairn, J., Duncan, D., Gray, L. M., Urquhart, G., Binnie, M., Byron, O. , Fothergill-Gilmore, L. A. and Price, N. C. (1998) Purification and characterization of pyruvate kinase from schizosaccharomyces pombe: evidence for an unusual quaternary structure. Protein Expression and Purification, 14(2), pp. 247-253. (doi:10.1006/prep.1998.0938) (PMID:9790887)

Nairn, J., Duncan, D., Gray, L. M., Urquhart, G., Binnie, M., Byron, O. , Fothergill-Gilmore, L. A. and Price, N. C. (1998) Purification and characterization of pyruvate kinase from schizosaccharomyces pombe: evidence for an unusual quaternary structure. Protein Expression and Purification, 14(2), pp. 247-253. (doi:10.1006/prep.1998.0938) (PMID:9790887)

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Abstract

Earlier attempts to purify and characterize nonrecombinant pyruvate kinase fromSchizosaccharomyces pombeproved difficult due to problems associated with the instability of the protein. The enzyme has been overexpressed inSaccharomyces cerevisiaestrain AH22, permitting studies to determine the conditions required to stabilize the enzyme during purification. RecombinantS. pombepyruvate kinase was purified by a combination of ion-exchange chromatography and gel filtration. The purified enzyme showed sigmoidal kinetics with respect to PEP; in the presence of FBP, the kinetics were restored to Michaelis–Menten behavior. With respect to ADP, the Hill coefficient was not affected by FBP. Determination of the molecular mass of the purified enzyme by ultracentrifugation showed that it behaved as a dimer–tetramer system with aKdof approximately 1 μM.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Byron, Professor Olwyn
Authors: Nairn, J., Duncan, D., Gray, L. M., Urquhart, G., Binnie, M., Byron, O., Fothergill-Gilmore, L. A., and Price, N. C.
College/School:College of Medical Veterinary and Life Sciences > School of Life Sciences
Journal Name:Protein Expression and Purification
Publisher:Elsevier
ISSN:1046-5928
ISSN (Online):1096-0279

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