The kinetic mechanism of phosphomevalonate kinase

Daniel Pilloff, Kristina Dabovic, Michael J. Romanowski, Jeffrey B. Bonanno, Mary Doherty, Stephen K. Burley, Thomas S. Leyh

Research output: Contribution to journalArticle

40 Citations (Scopus)

Abstract

Phosphomevalonate kinase catalyzes an essential step in the so-called mevalonate pathway, which appears to be the sole pathway for the biosynthesis of sterols and other isoprenoids in mammals and archea. Despite the well documented importance of this pathway in the cause and prevention of human disease and that it is the biosynthetic root of an enormous diverse class of metabolites, the mechanism of phosphomevalonate kinase from any organism is not yet well characterized. The first structure of a phosphomevalonate kinase from Streptococcus pneumoniae was solved recently. The enzyme exhibits an atypical P-loop that is a conserved defining feature of the GHMP kinase superfamily. In this study, the kinetic mechanism of the S. pneumoniae enzyme is characterized in the forward and reverse directions using a combination of classical initial-rate methods including alternate substrate inhibition using ADPβS. The inhibition patterns strongly support that in either direction the substrates bind randomly to the enzyme prior to chemistry, a random sequential bi-bi mechanism. The kinetic constants are as follows: kcat(forward) = 3.4 s-1, Ki(ATp) = 137 μM, Km(ATP) = 74 μM, Ki(pmev) = 7.7 μM, Km(pmev) = 4.2 μM; kcat(reverse) = 3.9 s-1, Ki(ADP) = 410 μM, Km(ADP) = 350 μM, Ki(ppmev) = 14 μM, Km(ppmev) = 12 μM, where pmev and ppmev represent phosphomevalonate and diphosphomevalonate, respectively.

Original languageEnglish (US)
Pages (from-to)4510-4515
Number of pages6
JournalJournal of Biological Chemistry
Volume278
Issue number7
DOIs
StatePublished - Feb 14 2003

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Streptococcus pneumoniae
Adenosine Diphosphate
Kinetics
Enzymes
Mevalonic Acid
Mammals
Biosynthesis
Archaea
Terpenes
Sterols
Substrates
Metabolites
Phosphotransferases
Adenosine Triphosphate
phosphomevalonate kinase
Direction compound
phosphomevalonic acid

ASJC Scopus subject areas

  • Biochemistry

Cite this

Pilloff, D., Dabovic, K., Romanowski, M. J., Bonanno, J. B., Doherty, M., Burley, S. K., & Leyh, T. S. (2003). The kinetic mechanism of phosphomevalonate kinase. Journal of Biological Chemistry, 278(7), 4510-4515. https://doi.org/10.1074/jbc.M210551200

The kinetic mechanism of phosphomevalonate kinase. / Pilloff, Daniel; Dabovic, Kristina; Romanowski, Michael J.; Bonanno, Jeffrey B.; Doherty, Mary; Burley, Stephen K.; Leyh, Thomas S.

In: Journal of Biological Chemistry, Vol. 278, No. 7, 14.02.2003, p. 4510-4515.

Research output: Contribution to journalArticle

Pilloff, D, Dabovic, K, Romanowski, MJ, Bonanno, JB, Doherty, M, Burley, SK & Leyh, TS 2003, 'The kinetic mechanism of phosphomevalonate kinase', Journal of Biological Chemistry, vol. 278, no. 7, pp. 4510-4515. https://doi.org/10.1074/jbc.M210551200
Pilloff D, Dabovic K, Romanowski MJ, Bonanno JB, Doherty M, Burley SK et al. The kinetic mechanism of phosphomevalonate kinase. Journal of Biological Chemistry. 2003 Feb 14;278(7):4510-4515. https://doi.org/10.1074/jbc.M210551200
Pilloff, Daniel ; Dabovic, Kristina ; Romanowski, Michael J. ; Bonanno, Jeffrey B. ; Doherty, Mary ; Burley, Stephen K. ; Leyh, Thomas S. / The kinetic mechanism of phosphomevalonate kinase. In: Journal of Biological Chemistry. 2003 ; Vol. 278, No. 7. pp. 4510-4515.
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