Magnesium acetate induces a conformational change in Escherichia coli primase.
Primase from Escherichia coli is a single-stranded DNA-dependent RNA polymerase. As such, it requires magnesium to carry out catalysis. Limited tryptic digestion was used to probe the conformations of primase as a function of magnesium acetate concentration. In the absence of magnesium, trypsin cleaved primase at three sites. Magnesium acetate induced a conformational change such that one of these sites became inaccessible to trypsin digestion and a new site became trypsin accessible. The conformational change was only induced by Mg(OAc)2 and not MnCl2, CaCl2, NaOAc or LiCl, indicating a clear magnesium acetate-dependent conformational change. The effect was slightly induced by MgSO4 and MgCl2. An allosteric binding model indicates that primase binds at least two magnesiums in a cooperative manner. The data were best fit to a two-state model in which one conformation had a high affinity for magnesium, KR = 83.4 M-1, and the other state had virtually no affinity.
Department of Chemistry, University of Nebraska-Lincoln 68588-0304, USA.
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Urlacher
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Griep
D000085:Acetates / Q000494:pharmacology*; D019342:Acetic Acid; D000495:Allosteric Site; D000595:Amino Acid Sequence; D002384:Catalysis; D019915:DNA Primase; D007700:Kinetics; D008956:Models, Chemical; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation / Q000187:drug effects; D012316:RNA Nucleotidyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014357:Trypsin
D000085:Acetates; D010446:Peptide Fragments; D019915:DNA Primase; D012316:RNA Nucleotidyltransferases; D014357:Trypsin; D019342:Acetic Acid