Abstract
This work first constructed the fusion protein NCRII-18 by fusing the restriction endonuclease Ecl18kI gene and part of the gene coding for the N-terminal domain of the endonuclease EcoRII. The fusion of the EcoRII N-terminal domain leads to a change in the properties of the recombinant protein. Unlike Ecl18kI, which made the basis of NCRII-18, the fusion protein predominantly recognizes the CCWGG sites, having lost the capability of interacting with the CCSGG sites. Experimental data support the hypothesis of a close evolutionary relationship between type IIE and IIP restriction endonucleases via a recombination between domains with active site structure and elements for recognition with domains responsible for recognition of DNA sequences.
Keywords:
DNA-protein interactions; domain organization; enterobacteria; fusion protein; restriction endonucleases.
© 2017 Federation of European Biochemical Societies.
MeSH terms
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Artificial Gene Fusion
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Binding Sites
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Catalytic Domain
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DNA Restriction Enzymes / chemistry
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DNA Restriction Enzymes / genetics
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DNA Restriction Enzymes / metabolism
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DNA, Bacterial / chemistry
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DNA, Bacterial / metabolism
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Deoxyribonucleases, Type II Site-Specific / chemistry
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Deoxyribonucleases, Type II Site-Specific / genetics*
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Deoxyribonucleases, Type II Site-Specific / metabolism
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Electrophoretic Mobility Shift Assay
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Escherichia coli / enzymology*
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Escherichia coli / genetics
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Escherichia coli Proteins / chemistry
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Escherichia coli Proteins / genetics*
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Escherichia coli Proteins / metabolism
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Evolution, Molecular*
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Gene Fusion*
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Models, Genetic*
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Nucleotide Motifs
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Peptide Fragments / chemistry
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Peptide Fragments / genetics
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Peptide Fragments / metabolism
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Protein Conformation
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Protein Engineering
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Recombinant Fusion Proteins / metabolism
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Recombination, Genetic
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Substrate Specificity
Substances
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DNA, Bacterial
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Escherichia coli Proteins
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Peptide Fragments
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Recombinant Fusion Proteins
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DNA Restriction Enzymes
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endodeoxyribonuclease Ecl18kI
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CCWGG-specific type II deoxyribonucleases
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Deoxyribonucleases, Type II Site-Specific