The zinc finger motif of Escherichia coli RecQ is implicated in both DNA binding and protein folding
The RecQ family of DNA helicases has been shown to be important for the maintenance of genomic integrity. Mutations in human RecQ genes lead to genomic instability and cancer. Several RecQ family of helicases contain a putative zinc finger motif of the C4 type at the C terminus that has been identified in the crystalline structure of RecQ helicase from Escherichia coli. To better understand the role of this motif in helicase from E. coli, we constructed a series of single mutations altering the conserved cysteines as well as other highly conserved residues. All of the resulting mutant proteins exhibited a high level of susceptibility to degradation, making functional analysis impossible. In contrast, a double mutant protein in which both cysteine residues Cys397 and Cys400 in the zinc finger motif were replaced by asparagine residues was purified to homogeneity. Slight local conformational changes were detected, but the rest of the mutant protein has a well defined tertiary structure. Furthermore, the mutant enzyme displayed ATP binding affinity similar to the wild-type enzyme but was severely impaired in DNA binding and in subsequent ATPase and helicase activities. These results revealed that the zinc finger binding motif is involved in maintaining the integrity of the whole protein as well as DNA binding. We also showed that the zinc atom is not essential to enzymatic activity.
| Medienart: |
Artikel |
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| Erscheinungsjahr: |
2004 |
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| Erschienen: |
2004 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:279 |
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| Enthalten in: |
The Journal of biological chemistry - 279(2004), 41 vom: 08. Okt., Seite 42794-802 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Liu, Jie Lin [VerfasserIn] |
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| Anmerkungen: |
Date Completed 24.11.2004 Date Revised 06.02.2021 published: Print-Electronic Citation Status MEDLINE |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM149737858 |
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| 100 | 1 | |a Liu, Jie Lin |e verfasserin |4 aut | |
| 245 | 1 | 4 | |a The zinc finger motif of Escherichia coli RecQ is implicated in both DNA binding and protein folding |
| 264 | 1 | |c 2004 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a ohne Hilfsmittel zu benutzen |b n |2 rdamedia | ||
| 338 | |a Band |b nc |2 rdacarrier | ||
| 500 | |a Date Completed 24.11.2004 | ||
| 500 | |a Date Revised 06.02.2021 | ||
| 500 | |a published: Print-Electronic | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a The RecQ family of DNA helicases has been shown to be important for the maintenance of genomic integrity. Mutations in human RecQ genes lead to genomic instability and cancer. Several RecQ family of helicases contain a putative zinc finger motif of the C4 type at the C terminus that has been identified in the crystalline structure of RecQ helicase from Escherichia coli. To better understand the role of this motif in helicase from E. coli, we constructed a series of single mutations altering the conserved cysteines as well as other highly conserved residues. All of the resulting mutant proteins exhibited a high level of susceptibility to degradation, making functional analysis impossible. In contrast, a double mutant protein in which both cysteine residues Cys397 and Cys400 in the zinc finger motif were replaced by asparagine residues was purified to homogeneity. Slight local conformational changes were detected, but the rest of the mutant protein has a well defined tertiary structure. Furthermore, the mutant enzyme displayed ATP binding affinity similar to the wild-type enzyme but was severely impaired in DNA binding and in subsequent ATPase and helicase activities. These results revealed that the zinc finger binding motif is involved in maintaining the integrity of the whole protein as well as DNA binding. We also showed that the zinc atom is not essential to enzymatic activity | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 7 | |a Chlorides |2 NLM | |
| 650 | 7 | |a Ions |2 NLM | |
| 650 | 7 | |a Zinc Compounds |2 NLM | |
| 650 | 7 | |a zinc chloride |2 NLM | |
| 650 | 7 | |a 86Q357L16B |2 NLM | |
| 650 | 7 | |a Adenosine Triphosphate |2 NLM | |
| 650 | 7 | |a 8L70Q75FXE |2 NLM | |
| 650 | 7 | |a DNA |2 NLM | |
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| 650 | 7 | |a Adenosine Triphosphatases |2 NLM | |
| 650 | 7 | |a EC 3.6.1.- |2 NLM | |
| 650 | 7 | |a RECQL protein, human |2 NLM | |
| 650 | 7 | |a EC 3.6.1.- |2 NLM | |
| 650 | 7 | |a RecQ protein, E coli |2 NLM | |
| 650 | 7 | |a EC 3.6.1.- |2 NLM | |
| 650 | 7 | |a DNA Helicases |2 NLM | |
| 650 | 7 | |a EC 3.6.4.- |2 NLM | |
| 650 | 7 | |a RecQ Helicases |2 NLM | |
| 650 | 7 | |a EC 3.6.4.12 |2 NLM | |
| 650 | 7 | |a Zinc |2 NLM | |
| 650 | 7 | |a J41CSQ7QDS |2 NLM | |
| 650 | 7 | |a Cysteine |2 NLM | |
| 650 | 7 | |a K848JZ4886 |2 NLM | |
| 700 | 1 | |a Rigolet, Pascal |e verfasserin |4 aut | |
| 700 | 1 | |a Dou, Shuo-Xing |e verfasserin |4 aut | |
| 700 | 1 | |a Wang, Peng-Ye |e verfasserin |4 aut | |
| 700 | 1 | |a Xi, Xu Guang |e verfasserin |4 aut | |
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