DOCSLIB.ORG

High-Resolution Structure of the E.Coli Recq Helicase Catalytic Core

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
High-Resolution Structure of the E.Coli Recq Helicase Catalytic Core The EMBO Journal Vol. 22 No. 19 pp. 4910±4921, 2003 High-resolution structure of the E.coli RecQ helicase catalytic core Douglas A.Bernstein, Morgan C.Zittel and repair replication forks that have stalled at sites of DNA James L.Keck1 damage (Courcelle et al., 1997; Courcelle and Hanawalt, 1999; Courcelle et al., 1999; 2003). In addition, E.coli Department of Biomolecular Chemistry, 550 Medical Science Center, 1300 University Avenue, University of Wisconsin Medical School, RecQ helps suppress illegitimate recombination in cells Madison, WI 53706-1532, USA (Hanada et al., 1997). Although the precise cellular substrates on which RecQ proteins act remain a mystery, 1Corresponding author e-mail: [email protected] several reactions catalyzed by E.coli RecQ have been reconstituted in vitro, including ATP-dependent DNA RecQ family helicases catalyze critical genome main- unwinding (Umezu et al., 1990), recombination initiation tenance reactions in bacterial and eukaryotic cells, with RecA and single-stranded (ss) DNA binding protein playing key roles in several DNA metabolic processes. (SSB) (Harmon and Kowalczykowski, 1998), and plasmid Mutations in recQ genes are linked to genome instabil- DNA catenation with topoisomerase III (Topo III) and ity and human disease. To de®ne the physical basis of SSB (Harmon et al., 1999). Coordinated activities with RecQ enzyme function, we have determined a 1.8 AÊ recombinases and topoisomerases, as well as DNA resolution crystal structure of the catalytic core of unwinding functions, are conserved in a number of Escherichia coli RecQ in its unbound form and a 2.5 AÊ eukaryotic RecQ proteins (reviewed in Wu and Hickson, resolution structure of the core bound to the ATP 2001), implying a functional homology among RecQ analog ATPgS. The RecQ core comprises four family members. conserved subdomains; two of these combine to Three diseases (Bloom's, Werner's, and Rothmund± form its helicase region, while the others form unex- Thompson syndromes) can be attributed to mutations of pected Zn2+-binding and winged-helix motifs. The human genes that encode homologs of E.coli RecQ (BLM, structures reveal the molecular basis of missense Ellis et al., 1995; WRN,Yuet al., 1996; RECQ4, Kitao mutations that cause Bloom's syndrome, a human et al., 1999). While each of these syndromes has distinct RecQ-associated disease. Finally, based on ®ndings clinical manifestations, they all share a predisposition to from the structures, we propose a mechanism for cancer. A common feature of these disorders is pro- RecQ activity that could explain its functional coordi- nounced genomic instability, which is characterized by nation with topoisomerase III. heightened levels of homologous recombination and/or Keywords: helicase/RecQ/structure/winged helix/Zn2+ chromosomal deletions (reviewed in Karow et al., 2000). binding Several disease-linked nonsense and frame-shift mutations in WRN, BLM, and RECQ4, as well as all known BLM missense mutations associated with Bloom's syndrome, map to regions that are conserved across the RecQ family Introduction of enzymes. RecQ DNA helicases are ubiquitous enzymes in bacteria Three regions conserved among bacterial and eukar- and eukaryotes that carry out important functions in yotic RecQ proteins have been identi®ed by sequence genome biology (Cobb et al., 2002; Wu and Hickson, analysis: helicase, RecQ-conserved (RecQ-Ct), and 2002). Like other DNA helicases, RecQ proteins are ATP- Helicase-and-RNaseD-C-terminal (HRDC) domains dependent molecular motors that unwind double-stranded (Figure 1A; Morozov et al., 1997). The helicase and (ds) DNA. Members of the RecQ family differ from other RecQ-Ct regions are the best conserved regions among helicases, however, in their unusual breadth of activities, RecQ proteins, with some RecQs lacking an identi®able including roles in DNA replication, recombination, and HRDC domain altogether. The RecQ family helicase repair. With their central activities in genome mainten- region contains sequence motifs that are broadly con- ance, it is not surprising that mutation of many known served among DNA unwinding enzymes (Gorbalenya and recQ genes leads to genome instability and, in the case of Koonin, 1993). These motifs link the energy of nucleoside three human recQ genes, to disease. Despite the import- triphosphate binding and hydrolysis to DNA unwinding. ance of RecQ proteins, the structural basis of their function The structure and function of the RecQ-Ct region is more has remained poorly de®ned. poorly understood since it appears to be unique to RecQ The founding RecQ family member was discovered in proteins. However, evidence indicates that this region is Escherichia coli where it functions in the RecF recombi- essential for RecQ function. Two disease-causing BLM nation pathway (Nakayama et al., 1984; Umezu et al., missense mutations map to the enzyme's RecQ-Ct region 1990). This pathway participates in homologous recombi- (Ellis et al., 1995; Foucault et al., 1997) and abolish BLM nation and repair of ultraviolet light-induced DNA damage ATPase and helicase activities in vitro (Bahr et al., 1998). (Kowalczykowski et al., 1994). RecQ and other proteins in Mutation and deletion analyses in the Saccharomyces the RecF pathway appear to catalyze reactions that help cerevisiae RecQ homolog (Sgs1) have also shown that this 4910 ã European Molecular Biology Organization Catalytic core of E.coli RecQ helicase Fig. 1. Structure of the E.coli RecQ catalytic core. (A) Schematic diagram of E.coli RecQ. Three conserved regions, helicase, RecQ-conserved (RecQ- Ct), and Helicase-and-RNaseD-C-terminal (HRDC) (Morozov et al., 1997), are labeled. The catalytic core of E.coli RecQ (RecQDC) includes only the helicase and RecQ-Ct regions, and comprises four apparent subdomains in the structure: residues 1±208 in red, 209±340 in blue, 341±406 in yellow, and 407±516 in green. (B) Sequence and secondary structure of RecQDC. Helices (boxes) and b-strands (arrows) are shown above the sequence and labeled sequentially. Color coding is the same as in (A). Conserved helicase motifs (motif 0, Bernstein and Keck, 2003; and motifs I-VI, Gorbalenya and Koonin, 1993) are labeled and enclosed in boxes. Residues that are invariant among 65 bacterial RecQ proteins are underlined, and residues that are invariant or highly conserved with a subset of eukaryotic RecQ proteins (human WRN, BLM, and S.cerevisiae Sgs1) are highlighted in purple or light-blue boxes, respectively. (C) Orthogonal views of a ribbon diagram of the crystal structure of RecQDC, color-coded as in (A). A bound Zn2+ ion is shown as a magenta sphere. 4911 D.A.Bernstein, M.C.Zittel and J.L.Keck region is essential for in vivo function (Mullen et al., 2000; (Janscak et al., 2003). Thus the question of the active Onoda et al., 2000; Ui et al., 2001). Together, the helicase oligomeric state of RecQ proteins remains unresolved. and RecQ-Ct regions form the core, conserved sequence To examine the physical basis of RecQ function, we that de®nes the RecQ family of enzymes. have determined two high-resolution X-ray crystal struc- Domain mapping experiments with E.coli RecQ have tures of the conserved catalytic core of E.coli RecQ. The demonstrated that the conserved helicase and RecQ-Ct core domain comprises four apparent subdomains; two of regions fold together to form a single 59 kDa structural the subdomains combine to form the helicase region and domain (RecQDC) (Bernstein and Keck, 2003). The the remaining two join to form the RecQ-Ct region of isolated RecQDC domain is active as a DNA-dependent RecQ. The RecQ-Ct subdomains include one that folds as ATPase and can unwind DNA with essentially the same a platform of helices that binds a Zn2+ ion using four speci®c activity as full-length E.coli RecQ, indicating that conserved cysteine residues and another that forms an it is the catalytic core of the enzyme (Bernstein and Keck, unexpected winged-helix (WH) subdomain. The WH 2003). The 9 kDa HRDC domain appears to modulate motif shares signi®cant structural similarity with other DNA binding activity of the catalytic core but is not DNA binding proteins, thus identifying a likely DNA necessary for enzyme function in vitro (Bernstein and binding site in E.coli RecQ. Based on the structures, we Keck, 2003). Similarly, an S.cerevisiae Sgs1 variant that propose a mechanism of action that could explain the lacks its C-terminal HRDC domain can complement the coordination of RecQ proteins with Topo III. Furthermore, increased DNA damage sensitivity phenotype of sgs1D our structures reveal the molecular basis of missense cells (Mullen et al., 2000; Mullen et al., 2001), implying mutations that cause Bloom's syndrome. Together, these that the HRDC domain is dispensable for at least some structures reveal the ®rst high-resolution images of the activities in vivo. In addition, a recent study has shown that enzymatic core conserved among RecQ family members. a catalytic core fragment of the human BLM protein that includes just its helicase and RecQ-Ct regions is active Results and discussion in vitro (Janscak et al., 2003). These ®ndings demonstrate that the evolutionarily conserved domain shared among Structure determination of the RecQDC domain RecQ proteins also forms the structural and catalytic core We ®rst attempted to crystallize full-length E.coli RecQ of the RecQ enzyme family. protein, but extensive crystallization trials yielded no Beyond low-resolution domain mapping, a number of crystals. This result could have been due to ¯exibility that other studies have revealed important structural features of has been observed between the catalytic core and HRDC RecQ helicases. First, sequence analysis places RecQ domains of E.coli RecQ (Bernstein and Keck, 2003) family members in the Superfamily 2 (SF2) group of leading to structural heterogeneity that would hinder enzymes (Gorbalenya and Koonin, 1993). While structur- crystallization.
Load more

Recommended publications
  • Structure and Function of the Human Recq DNA Helicases
    Zurich Open Repository and Archive University of Zurich Main Library Strickhofstrasse 39 CH-8057 Zurich www.zora.uzh.ch Year: 2005 Structure and function of the human RecQ DNA helicases Garcia, P L Posted at the Zurich Open Repository and Archive, University of Zurich ZORA URL: https://doi.org/10.5167/uzh-34420 Dissertation Published Version Originally published at: Garcia, P L. Structure and function of the human RecQ DNA helicases. 2005, University of Zurich, Faculty of Science. Structure and Function of the Human RecQ DNA Helicases Dissertation zur Erlangung der naturwissenschaftlichen Doktorw¨urde (Dr. sc. nat.) vorgelegt der Mathematisch-naturwissenschaftlichen Fakultat¨ der Universitat¨ Z ¨urich von Patrick L. Garcia aus Unterseen BE Promotionskomitee Prof. Dr. Josef Jiricny (Vorsitz) Prof. Dr. Ulrich H ¨ubscher Dr. Pavel Janscak (Leitung der Dissertation) Z ¨urich, 2005 For my parents ii Summary The RecQ DNA helicases are highly conserved from bacteria to man and are required for the maintenance of genomic stability. All unicellular organisms contain a single RecQ helicase, whereas the number of RecQ homologues in higher organisms can vary. Mu- tations in the genes encoding three of the five human members of the RecQ family give rise to autosomal recessive disorders called Bloom syndrome, Werner syndrome and Rothmund-Thomson syndrome. These diseases manifest commonly with genomic in- stability and a high predisposition to cancer. However, the genetic alterations vary as well as the types of tumours in these syndromes. Furthermore, distinct clinical features are observed, like short stature and immunodeficiency in Bloom syndrome patients or premature ageing in Werner Syndrome patients. Also, the biochemical features of the human RecQ-like DNA helicases are diverse, pointing to different roles in the mainte- nance of genomic stability.
    [Show full text]
  • 17 January 2001
    Running Title: DNA Recombination and Repair in the Archaea DNA Recombination and Repair in the Archaea Erica M. Seitz, Cynthia A. Haseltine, and Stephen C. Kowalczykowski* Sections of Microbiology and of Molecular and Cellular Biology Center for Genetics and Development University of California, Davis Davis, CA 95616-8665 * Corresponding author: Section of Microbiology One Shields Avenue Hutchison Hall University of California, Davis Davis, CA 95616-8665 Phone: (530)752-5938 Fax: (530)752-5939 email: [email protected] 1 Abstract The ability to repair DNA damage is crucial to all organisms. Much of what we learned about these processes was gained from studies carried out in Bacteria, especially in Escherichia coli, or Eucarya, particularly in the yeast Saccharomyces cerevisiae. The repair of DNA damage occurs by at least four different pathways: direct reversal of DNA damage, excision of damaged nucleotides (nucleotide excision repair or NER) or bases (base excision repair or BER), excision of misincorporated nucleotides (mismatch repair or MMR), and recombinational repair. Proteins involved in these processes have recently been identified in the third domain of life, the Archaea. Here we present a summary of DNA repair proteins in both the Bacteria and Eucarya, and discuss similarities and differences between these two domains and what is currently known in the Archaea. 2 I. Introduction DNA is subjected daily to considerable environmental and endogenous damage, which challenges both the integrity of the essential information that it contains and its ability to be transferred to future generations. All cells, however, are prepared to handle damage to the genome through an extensive DNA repair system, thus underscoring the importance of this process in cell survival.
    [Show full text]
  • The Role of Nucleotide Excision Repair in Restoring Replication Following UV-Induced Damage in Escherichia Coli
    Portland State University PDXScholar Dissertations and Theses Dissertations and Theses Summer 1-1-2012 The Role of Nucleotide Excision Repair in Restoring Replication Following UV-Induced Damage in Escherichia coli Kelley Nicole Newton Portland State University Follow this and additional works at: https://pdxscholar.library.pdx.edu/open_access_etds Part of the Biology Commons, and the Cell Biology Commons Let us know how access to this document benefits ou.y Recommended Citation Newton, Kelley Nicole, "The Role of Nucleotide Excision Repair in Restoring Replication Following UV- Induced Damage in Escherichia coli" (2012). Dissertations and Theses. Paper 767. https://doi.org/10.15760/etd.767 This Thesis is brought to you for free and open access. It has been accepted for inclusion in Dissertations and Theses by an authorized administrator of PDXScholar. Please contact us if we can make this document more accessible: [email protected]. The Role of Nucleotide Excision Repair in Restoring Replication Following UV-Induced Damage in Escherichia coli by Kelley Nicole Newton A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science in Biology Thesis Committee: Justin Courcelle, Chair Michael Bartlett Jeffrey Singer Portland State University 2012 ABSTRACT Following low levels of UV exposure, Escherichia coli cells deficient in nucleotide excision repair recover and synthesize DNA at near wild type levels, an observation that formed the basis of the post replication recombination repair model. In this study, we characterized the DNA synthesis that occurs following UV-irradiation in the absence of nucleotide excision repair and show that although this synthesis resumes at near wild type levels, it is coincident with a high degree of cell death.
    [Show full text]
  • Barbour-Thesis.Pdf (2.004Mb)
    Synthetically Lethal Interactions Classify Novel Genes in Postreplication Repair in Saccharomyces cerevisiae A thesis submitted to the College of Graduate Studies and Research in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy in the Department of Microbiology and Immunology University of Saskatchewan Leslie Barbour, B.Sc. © Copyright Leslie Barbour, February 2005. All rights reserved. Permission to Use In presenting this thesis in partial fulfillment of the requirements for a Postgraduate degree from the University of Saskatchewan, I agree that the libraries of this University may make it freely available for inspection. I further agree that permission for copying of this thesis in any manner, in whole or in part, for scholarly purposes may be granted by the professor or professors who supervised my thesis work or, in their absence, by the Head of the Department or the Dean of the College in which my thesis work was done. It is understood that any copying or publication or use of this thesis or parts thereof for financial gain shall not be allowed without my written permission. It is also understood that due recognition shall be given to me and to the University of Saskatchewan in any scholarly use which may be made of any material in my thesis. Requests for permission to copy or make other use of material in this thesis in whole or in part should be addressed to: Head of the Department of Microbiology and Immunology Health Sciences Building, 107 Wiggins Road University of Saskatchewan Saskatoon, SK Canada S7N 5E5 i Acknowledgments First I would like to thank my supervisor, Dr.
    [Show full text]
  • The Blm Helicase Literature Review
    UNIVERSITY OF CINCINNATI DATE: 3-22-03 I, Gregory T. Langland , hereby submit this as part of the requirements for the degree of: DOCTORATE OF PHILOSOPHY (Ph.D.) in: The Department of Molecular Genetics, Biochemistry and Microbiology of the College of Medicine It is entitled: INTERACTION BETWEEN THE BLM HELICASE AND THE DNA MISMATCH REPAIR PROTEIN, MLH1 Approved by: Joanna Groden Ph.D. Richard Wenstrup M.D. Jim Stringer Ph.D. Kathleen Dixon Ph.D. Peter Stambrook Ph.D. ii INTERACTION BETWEEN THE BLM HELICASE AND THE DNA MISMATCH REPAIR PROTEIN, MLH1 A dissertation submitted to the Division of Research and Advanced Studies of the University of Cincinnati in partial fulfillment of the requirements for the degree of DOCTORATE OF PHILOSOPHY (Ph.D.) In the Department of Molecular Genetics, Biochemistry & Microbiology of the College of Medicine 2002 by Gregory T. Langland B.S., University of Cincinnati, 1992 Committee Chair: Joanna Groden, Ph.D. iii `Abstract Bloom’s syndrome (BS) is a rare autosomal recessive disorder that greatly predisposes affected individuals to cancer. Such individuals also are small in size, sensitive to the sun, have immune dysfunction and gross genomic instability. The cytogenetics of BS cells have been extensively studied and have shown increased levels of homologous recombination, quadriradial formations, telomeric associations and chromosome breakage. The gene responsible for BS has been positionally cloned and and encodes a RecQ helicase family member with strand displacement activity that is dependent on ATP and Mg2+. In order to have a greater understanding of BLM helicase function in the cell in regards to DNA replication, recombination and repair, we identified protein- partners of BLM.
    [Show full text]
  • Initiation of Homologous Recombination
    February 3, 2004 Homologous Recombination by the RecBCD and RecF Pathways. Maria Spies and Stephen C. Kowalczykowski* Sections of Microbiology and of Molecular and Cellular Biology, Center for Genetics and Development, University of California, Davis, CA, 95616-8665 *CORRESPONDING AUTHOR: University of California, Davis, Section of Microbiology Center for Genetics and Development One Shields Ave. Davis, CA 95616-8665 E-mail: [email protected] Tel.: 530-752-5938 Fax: 530-752-5939 1 Abstract In all cells, genetic recombination is used to repair DNA breaks and, as a result, genetic information is exchanged between homologous DNA molecules. Discontinuities in DNA strands, specifically double-strand DNA breaks and single-strand DNA gaps, attract the enzymes responsible for the initiation of homologous recombination. In wild-type Escherichia coli, two distinct pathways are responsible for the repair of DNA by recombination: the RecBCD- and the RecF-pathways. The RecBCD-pathway is specific to recombination initiated at double-strand DNA breaks, whereas the RecF-pathway is primarily responsible for recombination initiated at single-strand DNA gaps, although it can also act at double-strand breaks. This review summarizes the biochemical mechanisms of homologous recombination in E. coli. 2 Recombinational repair of DNA damage. Homologous, or general, recombination is a crucial biological process that involves the pairing and transfer of strands between DNA molecules that share a region of significant sequence homology. Since its discovery in 1946 by Lederberg and Tatum (48), homologous recombination in bacteria was associated with the sexual process of conjugation and was viewed as an evolutionary mechanism both for shuffling the genome and for spreading favorable alleles.
    [Show full text]
  • The Werner Syndrome Gene Reviews the Werner Syndrome Gene the Molecular Basis of Recq Helicase- Deficiency Diseases
    The Werner syndrome gene Reviews The Werner syndrome gene the molecular basis of RecQ helicase- deficiency diseases Werner syndrome (WS) is an autosomal recessive genetic disorder that is manifested by genetic instability and premature onset of age-related diseases, including atherosclerosis and cancer. The gene that is mutated in WS cells (WRN) has been identified recently. Characterizations of the WRN gene product indicate that WRN encodes both a 39→59 DNA helicase, belonging to the Escherichia coli RecQ helicase family, and a 39→59 DNA exonuclease. Studies to define the molecular mechanism of WRN–DNA transactions are currently underway in many laboratories. Preliminary results indicate that WRN functions as a key factor in resolving aberrant DNA structures that arise from DNA metabolic processes such as replication, recombination and/or repair, to preserve the genetic integrity in cells. iscovered by Otto Werner in 1904 in a family dis- insights into the molecular bases of WS and other RecQ Dplaying symptoms similar to premature aging1, Werner helicase-deficiency diseases. syndrome (WS) is an uncommon autosomal recessive disorder characterized by early onset of age-related dis- WRN is a human DNA helicase homologous to eases including atherosclerosis, osteoporosis, type II diabetes E. coli RecQ mellitus, cataracts and rare soft-tissue sarcomas2. In cul- In 1992, the Werner syndrome gene (WRN) was mapped ture, cells from WS patients exhibit a shortened life-span3 to chromosome 8 at 8p12 (Ref. 21). The cDNA sequence and a prolonged S-phase of the cell cycle4. WS cells are of the entire gene was deciphered by positional cloning genetically unstable, as discovered initially by the finding and extensive DNA sequencing, and was reported in 1996 of nonclonal translocations, termed ‘variegated transloca- to encode a DNA helicase homologous to E.
    [Show full text]
  • From Resection to Resolution: Biochemical Investigation of Early and Late Steps of Homologous Recombination
    Zurich Open Repository and Archive University of Zurich Main Library Strickhofstrasse 39 CH-8057 Zurich www.zora.uzh.ch Year: 2016 From resection to resolution: biochemical investigation of early and late steps of homologous recombination Anand, Roopesh Abstract: Cells utilize two major pathways to repair DNA double strand breaks (DSB) including ho- mologous recombination (HR) and non-homologous end joining (NHEJ). DNA end resection of the 5’- terminated DNA at DSBs is the key event, which mechanistically determines the repair pathway choice by promoting HR while inhibiting NHEJ. Numerous studies have established the role of the MRE11- RAD50-NBS1 (MRN) complex and CtIP in the initiation of resection. However, the nucleolytic polarity of the only nuclease in the complex, MRE11, is the opposite (3’ to 5’) to the direction of resection (5’ to 3’). To solve this enigma, a so-called bidirectional resection model has been proposed. MRE11 was anticipated to make incision(s) close to the DSB by its endonucleolytic activity, which creates an entry site for its 3’ to 5’ exonuclease that proceeds back towards the DSB. Although this model solves the polarity paradox, the mechanistic understanding of such nicking activity by MRE11 was undefined. Here I show that CtIP stimulates the endonuclease activity of MRN on double-stranded DNA (dsDNA). Phos- phorylation of CtIP (pCtIP) at T847 is absolutely required for this stimulatory effect. The blocking of DNA ends, especially at the 5’ end, is required to observe this MRN-pCtIP activity. In agreement with the proposed model, the position of the first incision by MRN-pCtIP complex maps to [symbol about] 20 nucleotides away from the 5’ DNA end.
    [Show full text]
  • DNA Helicases and Human Disease
    22_DNARep_p.qxd 12/20/05 3:13 PM Page 1 22 DNA Helicases and Human Disease Ashwini S. Kamath-Loeb and Lawrence A. Loeb Joseph Gottstein Memorial Cancer Research Laboratory Departments of Pathology and Biochemistry University of Washington, Seattle, Washington 98195 Michael Fry Department of Biochemistry Rappaport Faculty of Medicine Technion–Israel Institute of Technology, Haifa, Israel HELICASES ARE MOTOR PROTEINS THAT UTILIZE THE ENERGY DERIVED from the hydrolysis of nucleoside triphosphates (NTP/dNTP) to disrupt hydrogen-bond interactions in double- or multi-stranded DNA and RNA. All known helicases are recognized to have at least two intrinsic en- zymatic activities: (1) NTP/dNTP-dependent nucleic acid unwinding and (2) DNA/RNA-dependent NTP/dNTP hydrolysis. Unwinding of DNA and RNA, with the generation of single-stranded nucleic acids, is essen- tial for numerous cellular transactions, including DNA replication, repair, and recombination, as well as RNA splicing, transcription, and transla- tion. Helicases therefore are ubiquitous, key players in several aspects of nucleic acid metabolism. The importance of helicases can be gleaned from the fact that, to date, 14 different DNA helicases have been identi- fied in bacteria, 15 in yeast, and 25 in human cells (for a compilation of known helicases, see Tuteja and Tuteja 2004). In addition, mutations in genes encoding DNA helicases have been demonstrated in several inher- ited human diseases. The fact that these diseases are rare also emphasizes the essentiality of DNA helicases in cellular metabolism. In this chapter we consider the different helicases, the proposed mechanisms for strand separation, their roles in cellular metabolism, and finally, the human dis- eases associated with mutations in specific DNA helicases.
    [Show full text]
  • LOSS of BLOOM SYNDROME PROTEIN CAUSES DESTABILIZATION of GENOMIC ARCHITECTURE and IS COMPLEMENTED by ECTOPIC EXPRESSION of Escherichia Coli Recg in HUMAN CELLS
    University of Kentucky UKnowledge University of Kentucky Doctoral Dissertations Graduate School 2011 LOSS OF BLOOM SYNDROME PROTEIN CAUSES DESTABILIZATION OF GENOMIC ARCHITECTURE AND IS COMPLEMENTED BY ECTOPIC EXPRESSION OF Escherichia coli RecG IN HUMAN CELLS Michael Wayne Killen University of Kentucky, [email protected] Right click to open a feedback form in a new tab to let us know how this document benefits ou.y Recommended Citation Killen, Michael Wayne, "LOSS OF BLOOM SYNDROME PROTEIN CAUSES DESTABILIZATION OF GENOMIC ARCHITECTURE AND IS COMPLEMENTED BY ECTOPIC EXPRESSION OF Escherichia coli RecG IN HUMAN CELLS" (2011). University of Kentucky Doctoral Dissertations. 201. https://uknowledge.uky.edu/gradschool_diss/201 This Dissertation is brought to you for free and open access by the Graduate School at UKnowledge. It has been accepted for inclusion in University of Kentucky Doctoral Dissertations by an authorized administrator of UKnowledge. For more information, please contact [email protected]. ABSTRACT OF DISSERTATION Michael Wayne Killen The Graduate School University of Kentucky 2011 LOSS OF BLOOM SYNDROME PROTEIN CAUSES DESTABILIZATION OF GENOMIC ARCHITECTURE AND IS COMPLEMENTED BY ECTOPIC EXPRESSION OF Escherichia coli RecG IN HUMAN CELLS ABSTRACT OF DISSERTATION A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the College of Medicine, Department of Microbiology, Immunology, and Molecular Genetics at the University of Kentucky By Michael Wayne Killen
    [Show full text]
  • Structural Framework for the Mechanism of Archaeal Exosomes in RNA Processing
    Dissertation zur Erlangung des Doktorgrades der Fakultät für Chemie und Pharmazie der Ludwig-Maximilians-Universität München Structural Framework for the Mechanism of Archaeal Exosomes in RNA processing Structural Insights into DNA Duplex Separation by the Archaeal Superfamily 2 Helicase Hel308 Katharina Büttner aus Wolfratshausen 2007 Erklärung Diese Dissertation wurde im Sinne von § 13 Abs. 3 bzw. 4 der Promotionsordnung vom 29. Januar 1998 von Herrn Prof. Dr. Karl-Peter Hopfner betreut. Ehrenwörtliche Versicherung Diese Dissertation wurde selbständig, ohne unerlaubte Hilfe erarbeitet. München, am 24. April 2007 .................................................... (Katharina Büttner) Dissertation eingereicht am 24. April 2007 1. Gutachter Herr Prof. Dr. Karl-Peter Hopfner 2. Gutachter Herr Prof. Dr. Patrick Cramer Mündliche Prüfung am 03. Juli 2007 The presented thesis was prepared in the time from December 2003 to March 2007 in the laboratory of Professor Dr. Karl-Peter Hopfner at the Gene Center of the Ludwig-Maximilians-University of Munich (LMU). Parts of this PhD thesis have been published: Büttner K., Nehring S., and Hopfner K.P., (2007). Structural basis for DNA duplex separation by a Superfamily 2 Helicase. NSMB 2007, Jul;14(7):647-52. Büttner K. and Hopfner K.P., (2006). Das Exosom. Bioforum 2006; 29:19-21. Büttner K., Wenig K., and Hopfner K.P., (2006). The exosome: a macromolecular cage for controlled RNA degradation. Mol. Microbiology 2006;61:1372-1379. Büttner K., Wenig K.. and Hopfner K.P., (2005). Structural Framework for the Mechanism of Archaeal Exosomes in RNA Processing. Mol Cell. 2005 Nov 11;20(3):461-71. Other publications: Karcher A., Büttner K., Märtens B., Jansen R.P.
    [Show full text]
  • Homologous Recombination by the Recbcd and Recf Pathways
    The Bactertal Chromosome Edited by N.Patrick Higgins 2005 ASM Press, Washington, D.C. Chapter 21 Homologous Recombination by the RecBCD and RecF Pathways RECOMBINATIONAL REPAIR specific for a particular DNA lesion (see reference 28 OF DNA DAMAGE for a review). These repair systems use the intact com- plementary strand as a template to restore the dam- Homologous, or general, recombination is a cru- aged DNkmolecule to its original state. Occasionally cial biological process that involves the paring and both strands of the double-stranded DNA (dsDNA) transfer of strands between DNA molecules that share can be broken opposite to each other, resulting in a a region of significant sequence homology. After its double-strand break (DSB). DSBs can be produced, discovery in 1946 by Lederberg and Tatum (48), ho- for example, as a direct consequence of ionizing ra- mologous recombination in bacteria was associated diation (Fig. 1A). However, the bulk of the DSBs with the sexual process of conjugation and was viewed in bacteria are generated indirectly as the result of as an evolutionary mechanism both for shuffling the DNA replication through an unrepaired break in just genome and for spreading favorable alleles. However, a single strand of DNA (Fig. 1B). Replication of DNA more recently, a more immediate function of homol- containing a single-strand nick or a gap in the leading ogous recombination has been recognized: namely, it strand results in the dissociation of the DNA poly- is a mechanism for the maintenance of chromosomal merase holoenzyme complex and the generation of integrity that acts to repair DNA lesions, both double- one blunt DSB (Fig.
    [Show full text]