title: Temperature effects on the allosteric transition of ATP sulfurylase from Penicillium chrysogenum.
abstract: The effects of temperature on the initial velocity kinetics of allosteric ATP sulfurylase from Penicillium chrysogenum were measured. The experiments were prompted by the structural similarity between the C-terminal regulatory domain of fungal ATP sulfurylase and fungal APS kinase, a homodimer that undergoes a temperature-dependent, reversible dissociation of subunits over a narrow temperature range. Wild-type ATP sulfurylase yielded hyperbolic velocity curves between 18 and 30 degrees C. Increasing the assay temperature above 30 degrees C at a constant pH of 8.0 increased the cooperativity of the velocity curves. Hill coefficients (n(H)) up to 1.8 were observed at 42 degrees C. The bireactant kinetics at 42 degrees C were the same as those observed at 30 degrees C in the presence of PAPS, the allosteric inhibitor. In contrast, yeast ATP sulfurylase yielded hyperbolic plots at 42 degrees C. The P. chrysogenum mutant enzyme, C509S, which is intrinsically cooperative (n(H) = 1.8) at 30 degrees C, became more cooperative as the temperature was increased yielding n(H) values up to 2.9 at 42 degrees C. As the temperature was decreased, the cooperativity of C509S decreased; n(H) was 1.0 at 18 degrees C. The cumulative results indicate that increasing the temperature increases the allosteric constant, L, i.e., promotes a shift in the base-level distribution of enzyme molecules from the high MgATP affinity R state toward the low MgATP affinity T state. As a result, the enzyme displays a true "temperature optimum" at subsaturating MgATP. The reversible temperature-dependent transitions of fungal ATP sulfurylase and APS kinase may play a role in energy conservation at high temperatures where the organism can survive but not grow optimally.
authors.affiliation: Section of Molecular and Cellular Biology, University of California, One Shields Avenue, Davis, California, 95616, USA.
authors.forename: D C
authors.initials: DC
authors.lastname: Medina
authors.forename: E
authors.initials: E
authors.lastname: Hanna
authors.forename: I J
authors.initials: IJ
authors.lastname: MacRae
authors.forename: A J
authors.initials: AJ
authors.lastname: Fisher
authors.forename: I H
authors.initials: IH
authors.lastname: Segel
mesh_terms: D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D019281:Dimerization; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010408:Penicillium chrysogenum / Q000201:enzymology* / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013430:Sulfate Adenylyltransferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013696:Temperature
chemical_list: D017853:Phosphotransferases (Alcohol Group Acceptor); C019617:adenylylsulfate kinase; D013430:Sulfate Adenylyltransferase