Abstract
Caulobacter crescentus relies on DNA methylation by the cell cycle-regulated methyltransferase (CcrM) in addition to key transcription factors to control the cell cycle and direct cellular differentiation. CcrM is shown here to efficiently methylate its cognate recognition site 5'-GANTC-3' in single-stranded and hemimethylated double-stranded DNA. We report the Km, kcat, kmethylation, and Kd for single-stranded and hemimethylated substrates, revealing discrimination of 107-fold for noncognate sequences. The enzyme also shows a similar discrimination against single-stranded RNA. Two independent assays clearly show that CcrM is highly processive with single-stranded and hemimethylated DNA. Collectively, the data provide evidence that CcrM and other DNA-modifying enzymes may use a new mechanism to recognize DNA in a key epigenetic process.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Caulobacter crescentus / cytology
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Caulobacter crescentus / enzymology*
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Cell Cycle
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Coenzymes / metabolism
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DNA / chemistry
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DNA / metabolism*
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DNA Methylation*
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DNA, Single-Stranded / chemistry
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DNA, Single-Stranded / metabolism*
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Electrophoretic Mobility Shift Assay
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Fluoresceins / analysis
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Fluorescent Dyes / analysis
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Kinetics
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Nucleotide Motifs
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RNA / chemistry
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RNA / metabolism
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Recombinant Fusion Proteins / metabolism
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Recombinant Proteins / metabolism
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S-Adenosylmethionine / metabolism
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Site-Specific DNA-Methyltransferase (Adenine-Specific) / genetics
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Site-Specific DNA-Methyltransferase (Adenine-Specific) / metabolism*
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Substrate Specificity
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Thermodynamics
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Tritium
Substances
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Coenzymes
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DNA, Single-Stranded
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Fluoresceins
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Fluorescent Dyes
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Recombinant Fusion Proteins
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Recombinant Proteins
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Tritium
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RNA
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S-Adenosylmethionine
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DNA
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DNA modification methylase CcrM
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DNA modification methylase HindII
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Site-Specific DNA-Methyltransferase (Adenine-Specific)