title: ATP hydrolysis and DNA binding confer thermostability on the MCM helicase.
abstract: The minichromosome maintenance (MCM) helicase is the replicative helicase in archaea. The enzyme utilizes the energy derived from ATP hydrolysis to translocate along one strand of the DNA and unwind the complementary strand. Here, the effect of DNA and ATP on the thermostability of the Methanothermobacter thermautotrophicus MCM protein was determined by differential scanning calorimetry. The MCM protein shows a single thermal transition at 67 degrees C. The stability is dramatically altered with the appearance of a second thermal transition up to 10 degrees C higher in the presence of DNA and either ATP or ADP-AlF(4)(-), a transition-state analogue of ATP, bound to MCM. In the presence of DNA and ADP or the nonhydrolyzable ATP analogues ATPgammaS and AMP-PNP, however, only a single thermal transition is observed at temperatures slightly higher than the transition temperature of MCM alone. Thus, the results suggest that ATP hydrolysis proceeds through a transition state that decouples an interaction between the N-terminal DNA binding domain and the C-terminal catalytic domain in the presence of DNA.
authors.affiliation: University of Maryland Biotechnology Institute, Center for Advanced Research in Biotechnology, Rockville, Maryland 20850, USA.
authors.forename: Nozomi
authors.initials: N
authors.lastname: Sakakibara
authors.forename: Frederick P
authors.initials: FP
authors.lastname: Schwarz
authors.forename: Zvi
authors.initials: Z
authors.lastname: Kelman
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000378:metabolism; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004265:DNA Helicases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004795:Enzyme Stability; D006358:Hot Temperature; D006868:Hydrolysis; D017013:Methanobacteriaceae / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary
chemical_list: D019843:Archaeal Proteins; D004247:DNA; D000251:Adenosine Triphosphatases; D004265:DNA Helicases; C403162:MCM protein, Methanobacterium thermoautotrophicum