Kinetic and Stability Properties of Penicillium chrysogenum ATP Sulfurylase Missing the C-terminal Regulatory Domain

Eissa Hanna, Kit Fai Ng, Ian J. MacRae, Christopher J. Bley, Andrew J Fisher, Irwin H. Segel

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13 Scopus citations


ATP sulfurylase from Penicillium chrysogenum is a homohexameric enzyme that is subject to allosteric inhibition by 3′-phosphoadenosine 5′-phosphosulfate. In contrast to the wild type enzyme, recombinant ATP sulfurylase lacking the C-terminal allosteric domain was monomeric and noncooperative. All kcat values were decreased (the adenosine 5′-phosphosulfate (adenylylsulfate) (APS) synthesis reaction to 17% of the wild type value). Additionally, the Michaelis constants for MgATP and sulfate (or molybdate), the dissociation constant of E·APS, and the monovalent oxyanion dissociation constants of dead end E·MgATP· oxyanion complexes were all increased. APS release (the k6 step) was rate-limiting in the wild type enzyme. Without the C-terminal domain, the composite k5 step (isomerization of the central complex and MgPP i release) became rate-limiting. The cumulative results indicate that besides (a) serving as a receptor for the allosteric inhibitor, the C-terminal domain (b) stabilizes the hexameric structure and indirectly, individual subunits. Additionally, (c) the domain interacts with and perfects the catalytic site such that one or more steps following the formation of the binary E·MgATP and E·SO4 2- complexes and preceding the release of MgPPi are optimized. The more negative entropy of activation of the truncated enzyme for APS synthesis is consistent with a role of the C-terminal domain in promoting the effective orientation of MgATP and sulfate at the active site.

Original languageEnglish (US)
Pages (from-to)4415-4424
Number of pages10
JournalJournal of Biological Chemistry
Issue number6
StatePublished - Feb 6 2004

ASJC Scopus subject areas

  • Biochemistry


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