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 |
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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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