Kumar, S and Bangru, S and Kumar, R and Rao, DN (2020) Promiscuous DNA cleavage by HpyAII endonuclease is modulated by the HNH catalytic residues. In: Bioscience Reports, 40 (9).
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Abstract
Helicobacter pylori is a carcinogenic bacterium that is responsible for 5.5 of all human gastric cancers. H. pylori codes for an unusually large number of restriction-modification (R-M) systems and several of them are strain-specific and phase-variable. HpyAII is a novel Type IIs phase-variable restriction endonuclease present in 26695 strain of H. pylori. We show that HpyAII prefers two-site substrates over one-site substrates for maximal cleavage activity. HpyAII is less stringent in metal ion requirement and shows higher cleavage activity with Ni2+ over Mg2+. Mutational analysis of the putative residues of the HNH motif of HpyAII confirms that the protein has an active HNH site for the cleavage of DNA. However, mutation of the first Histidine residue of the HNH motif to Alanine does not abolish the enzymatic activity, but instead causes loss of fidelity compared with wildtype HpyAII. Previous studies have shown that mutation of the first Histidine residue of the HNH motif of all other known HNH motif motif-containing enzymes completely abolishes enzymatic activity. We found, in the case of HpyAII, mutation of an active site residue leads to the loss of endonuclease fidelity. The present study provides further insights into the evolution of restriction enzymes. ©2020 The Author(s).
| Item Type: | Journal Article |
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| Publication: | Bioscience Reports |
| Publisher: | Portland Press Ltd |
| Additional Information: | The copyright for this article belongs to The Author(s). |
| Keywords: | endonuclease; HpyAII endonuclease; magnesium; nickel; unclassified drug; bacterial protein; divalent cation; type II site specific deoxyribonuclease, Article; bacterial cell; bacterial strain; catalysis; circular dichroism; controlled study; DNA cleavage; DNA cleavage assay; enzyme active site; enzyme activity; enzyme structure; enzyme substrate; Escherichia coli; Helicobacter pylori; mass spectrometry; molecular cloning; mutational analysis; nonhuman; open reading frame; peptide mass fingerprinting; protein motif; sequence alignment; sequence analysis; site directed mutagenesis; binding site; coenzyme; DNA cleavage; enzyme specificity; enzymology; genetics; metabolism, Bacterial Proteins; Binding Sites; Catalytic Domain; Cations, Divalent; Coenzymes; Deoxyribonucleases, Type II Site-Specific; DNA Cleavage; Helicobacter pylori; Magnesium; Mutagenesis, Site-Directed; Nickel; Substrate Specificity |
| Department/Centre: | Division of Biological Sciences > Biochemistry |
| Date Deposited: | 13 Jan 2023 05:39 |
| Last Modified: | 13 Jan 2023 05:39 |
| URI: | https://eprints.iisc.ac.in/id/eprint/79093 |
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