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Bylund, Göran
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Publications (10 of 12) Show all publications
ter Beek, J., Parkash, V., Bylund, G., Osterman, P., Sauer-Eriksson, A. E. & Johansson, E. (2019). Structural evidence for an essential Fe–S cluster in the catalytic core domain of DNA polymerase ϵ. Nucleic Acids Research, 47(11), 5712-5722
Open this publication in new window or tab >>Structural evidence for an essential Fe–S cluster in the catalytic core domain of DNA polymerase ϵ
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2019 (English)In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 47, no 11, p. 5712-5722Article in journal (Refereed) Published
Abstract [en]

DNA polymerase ϵ (Pol ϵ), the major leading-strand DNA polymerase in eukaryotes, has a catalytic subunit (Pol2) and three non-catalytic subunits. The N-terminal half of Pol2 (Pol2CORE) exhibits both polymerase and exonuclease activity. It has been suggested that both the non-catalytic C-terminal domain of Pol2 (with the two cysteine motifs CysA and CysB) and Pol2CORE (with the CysX cysteine motif) are likely to coordinate an Fe–S cluster. Here, we present two new crystal structures of Pol2CORE with an Fe–S cluster bound to the CysX motif, supported by an anomalous signal at that position. Furthermore we show that purified four-subunit Pol ϵ, Pol ϵ CysAMUT (C2111S/C2133S), and Pol ϵ CysBMUT (C2167S/C2181S) all have an Fe–S cluster that is not present in Pol ϵ CysXMUT (C665S/C668S). Pol ϵ CysAMUT and Pol ϵ CysBMUT behave similarly to wild-type Pol ϵ in in vitro assays, but Pol ϵ CysXMUT has severely compromised DNA polymerase activity that is not the result of an excessive exonuclease activity. Tetrad analyses show that haploid yeast strains carrying CysXMUT are inviable. In conclusion, Pol ϵ has a single Fe–S cluster bound at the base of the P-domain, and this Fe–S cluster is essential for cell viability and polymerase activity.

Place, publisher, year, edition, pages
Oxford University Press, 2019
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-161925 (URN)10.1093/nar/gkz248 (DOI)000475702000027 ()30968138 (PubMedID)2-s2.0-85068487970 (Scopus ID)
Available from: 2019-08-06 Created: 2019-08-06 Last updated: 2019-08-06Bibliographically approved
Bugaytsova, J. A., Björnham, O., Chernov, Y. A., Gideonsson, P., Henriksson, S., Mendez, M., . . . Boren, T. (2017). Helicobacter pylori Adapts to Chronic Infection and Gastric Disease via pH-Responsive BabA-Mediated Adherence. Cell Host and Microbe, 21(3), 376-389
Open this publication in new window or tab >>Helicobacter pylori Adapts to Chronic Infection and Gastric Disease via pH-Responsive BabA-Mediated Adherence
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2017 (English)In: Cell Host and Microbe, ISSN 1931-3128, E-ISSN 1934-6069, Vol. 21, no 3, p. 376-389Article in journal (Refereed) Published
Abstract [en]

The BabA adhesin mediates high-affinity binding of Helicobacter pylori to the ABO blood group antigen-glycosylated gastric mucosa. Here we show that BabA is acid responsive-binding is reduced at low pH and restored by acid neutralization. Acid responsiveness differs among strains; often correlates with different intragastric regions and evolves during chronic infection and disease progression; and depends on pH sensor sequences in BabA and on pH reversible formation of high-affinity binding BabA multimers. We propose that BabA's extraordinary reversible acid responsiveness enables tight mucosal bacterial adherence while also allowing an effective escape from epithelial cells and mucus that are shed into the acidic bactericidal lumen and that bio-selection and changes in BabA binding properties through mutation and recombination with babA-related genes are selected by differences among individuals and by changes in gastric acidity over time. These processes generate diverse H. pylori subpopulations, in which BabA's adaptive evolution contributes to H. pylori persistence and overt gastric disease.

Place, publisher, year, edition, pages
CELL PRESS, 2017
National Category
Microbiology in the medical area Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-132788 (URN)10.1016/j.chom.2017.02.013 (DOI)000396375600023 ()28279347 (PubMedID)
Available from: 2017-05-11 Created: 2017-05-11 Last updated: 2019-05-24Bibliographically approved
Yousefzadeh, M. J., Wyatt, D. W., Takata, K., Mu, Y., Hensley, S. C., Tomida, J., . . . Wood, R. D. (2015). Mammalian POLQ, Chromosome Stability and DNA Double-Strand Break Repair. Environmental and Molecular Mutagenesis, 56, S48-S48
Open this publication in new window or tab >>Mammalian POLQ, Chromosome Stability and DNA Double-Strand Break Repair
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2015 (English)In: Environmental and Molecular Mutagenesis, ISSN 0893-6692, E-ISSN 1098-2280, Vol. 56, p. S48-S48Article in journal, Meeting abstract (Other academic) Published
National Category
Pharmacology and Toxicology Medical Genetics
Identifiers
urn:nbn:se:umu:diva-108466 (URN)000360226400070 ()
Available from: 2015-09-14 Created: 2015-09-11 Last updated: 2018-06-07Bibliographically approved
Ganai, R. A., Bylund, G. & Johansson, E. (2015). Switching between polymerase and exonuclease sites in DNA polymerase ε. Nucleic Acids Research, 43(2), 932-942
Open this publication in new window or tab >>Switching between polymerase and exonuclease sites in DNA polymerase ε
2015 (English)In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 43, no 2, p. 932-942Article in journal (Refereed) Published
Abstract [en]

The balance between exonuclease and polymerase activities promotes DNA synthesis over degradation when nucleotides are correctly added to the new strand by replicative B-family polymerases. Misincorporations shift the balance toward the exonuclease site, and the balance tips back in favor of DNA synthesis when the incorrect nucleotides have been removed. Most B-family DNA polymerases have an extended β-hairpin loop that appears to be important for switching from the exonuclease site to the polymerase site, a process that affects fidelity of the DNA polymerase. Here, we show that DNA polymerase ε can switch between the polymerase site and exonuclease site in a processive manner despite the absence of an extended β-hairpin loop. K967 and R988 are two conserved amino acids in the palm and thumb domain that interact with bases on the primer strand in the minor groove at positions n−2 and n−4/n−5, respectively. DNA polymerase ε depends on both K967 and R988 to stabilize the 3′-terminus of the DNA within the polymerase site and on R988 to processively switch between the exonuclease and polymerase sites. Based on a structural alignment with DNA polymerase δ, we propose that arginines corresponding to R988 might have a similar function in other B-family polymerases.

Place, publisher, year, edition, pages
Oxford University Press, 2015
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-97693 (URN)10.1093/nar/gku1353 (DOI)000350209000027 ()25550436 (PubMedID)
Available from: 2015-01-08 Created: 2015-01-05 Last updated: 2018-06-07Bibliographically approved
Yousefzadeh, M. J., Wyatt, D. W., Takata, K.-I., Mu, Y., Hensley, S. C., Tomida, J., . . . Wood, R. D. (2014). Mechanism of suppression of chromosomal instability by DNA polymerase POLQ. PLOS Genetics, 10(10), e1004654
Open this publication in new window or tab >>Mechanism of suppression of chromosomal instability by DNA polymerase POLQ
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2014 (English)In: PLOS Genetics, ISSN 1553-7390, E-ISSN 1553-7404, Vol. 10, no 10, p. e1004654-Article in journal (Refereed) Published
Abstract [en]

Although a defect in the DNA polymerase POLQ leads to ionizing radiation sensitivity in mammalian cells, the relevant enzymatic pathway has not been identified. Here we define the specific mechanism by which POLQ restricts harmful DNA instability. Our experiments show that Polq-null murine cells are selectively hypersensitive to DNA strand breaking agents, and that damage resistance requires the DNA polymerase activity of POLQ. Using a DNA break end joining assay in cells, we monitored repair of DNA ends with long 3' single-stranded overhangs. End joining events retaining much of the overhang were dependent on POLQ, and independent of Ku70. To analyze the repair function in more detail, we examined immunoglobulin class switch joining between DNA segments in antibody genes. POLQ participates in end joining of a DNA break during immunoglobulin class-switching, producing insertions of base pairs at the joins with homology to IgH switch-region sequences. Biochemical experiments with purified human POLQ protein revealed the mechanism generating the insertions during DNA end joining, relying on the unique ability of POLQ to extend DNA from minimally paired primers. DNA breaks at the IgH locus can sometimes join with breaks in Myc, creating a chromosome translocation. We found a marked increase in Myc/IgH translocations in Polq-defective mice, showing that POLQ suppresses genomic instability and genome rearrangements originating at DNA double-strand breaks. This work clearly defines a role and mechanism for mammalian POLQ in an alternative end joining pathway that suppresses the formation of chromosomal translocations. Our findings depart from the prevailing view that alternative end joining processes are generically translocation-prone.

Place, publisher, year, edition, pages
Public library science, 2014
National Category
Other Basic Medicine Medical Genetics Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-94748 (URN)10.1371/journal.pgen.1004654 (DOI)000344650700030 ()25275444 (PubMedID)
Available from: 2014-10-16 Created: 2014-10-16 Last updated: 2018-06-07Bibliographically approved
Hogg, M., Osterman, P., Bylund, G., Ganai, R. A., Lundström, E.-B., Sauer-Eriksson, E. & Johansson, E. (2014). Structural basis for processive DNA synthesis by yeast DNA polymerase ε. Nature Structural & Molecular Biology, 21(1), 49-56
Open this publication in new window or tab >>Structural basis for processive DNA synthesis by yeast DNA polymerase ε
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2014 (English)In: Nature Structural & Molecular Biology, ISSN 1545-9993, E-ISSN 1545-9985, Vol. 21, no 1, p. 49-56Article in journal (Refereed) Published
Abstract [en]

DNA polymerase ε (Pol ε) is a high-fidelity polymerase that has been shown to participate in leading-strand synthesis during DNA replication in eukaryotic cells. We present here a ternary structure of the catalytic core of Pol ε (142 kDa) from Saccharomyces cerevisiae in complex with DNA and an incoming nucleotide. This structure provides information about the selection of the correct nucleotide and the positions of amino acids that might be critical for proofreading activity. Pol ε has the highest fidelity among B-family polymerases despite the absence of an extended b-hairpin loop that is required for high-fidelity replication by other B-family polymerases. Moreover, the catalytic core has a new domain that allows Pol ε to encircle the nascent doublestranded DNA. Altogether, the structure provides an explanation for the high processivity and high fidelity of leading-strand DNA synthesis in eukaryotes

Place, publisher, year, edition, pages
Nature Publishing Group, 2014
National Category
Biochemistry and Molecular Biology
Research subject
Biochemistry
Identifiers
urn:nbn:se:umu:diva-97700 (URN)10.1038/nsmb.2712 (DOI)000329290700014 ()24292646 (PubMedID)
Available from: 2015-01-08 Created: 2015-01-05 Last updated: 2018-06-07Bibliographically approved
Bylund, G. O., Nord, S., Lövgren, J. M. & Wikström, P. M. (2011). Alterations in the β flap and β' dock domains of the RNA polymerase abolish NusA-mediated feedback regulation of the metY-nusA-infB operon. Journal of Bacteriology, 193(16), 4113-4122
Open this publication in new window or tab >>Alterations in the β flap and β' dock domains of the RNA polymerase abolish NusA-mediated feedback regulation of the metY-nusA-infB operon
2011 (English)In: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 193, no 16, p. 4113-4122Article in journal (Refereed) Published
Abstract [en]

The RimM protein in Escherichia coli is important for the in vivo maturation of 30S ribosomal subunits and a ΔrimM mutant grows poorly due to assembly and translational defects. These deficiencies are suppressed partially by mutations that increase the synthesis of another assembly protein, RbfA, encoded by the metY-nusA-infB operon. Among these suppressors are mutations in nusA that impair the NusA-mediated negative-feedback regulation at internal intrinsic transcriptional terminators of the metY-nusA-infB operon. We describe here the isolation of two new mutations, one in rpoB and one in rpoC (encoding the β and β' subunits of the RNA polymerase, respectively), that increase the synthesis of RbfA by preventing NusA from stimulating termination at the internal intrinsic transcriptional terminators of the metY-nusA-infB operon. The rpoB2063 mutation changed the isoleucine in position 905 of the β flap-tip helix to a serine, while the rpoC2064 mutation duplicated positions 415 to 416 (valine-isoleucine) at the base of the β' dock domain. These findings support previously published in vitro results, which have suggested that the β flap-tip helix and β' dock domain at either side of the RNA exit tunnel mediate the binding to NusA during transcriptional pausing and termination.

Keywords
ribosome maturation protein; escherichia-coli operon; termination factor-rho; messenger-rna; polynucleotide phosphorylase; transcription elongation; nucleotide-sequence; level expression; gene; rimm
National Category
Natural Sciences Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-61270 (URN)10.1128/JB.00196-11 (DOI)21685293 (PubMedID)
Available from: 2012-11-07 Created: 2012-11-07 Last updated: 2018-06-08Bibliographically approved
Fei, Y. Y., Schmidt, A., Bylund, G., Johansson, D. X., Henriksson, S., Lebrilla, C., . . . Zhu, X. D. (2011). Use of real-time, label-free analysis in revealing low-affinity binding to blood group antigens by Helicobacter pylori. Analytical Chemistry, 83(16), 6336-6341
Open this publication in new window or tab >>Use of real-time, label-free analysis in revealing low-affinity binding to blood group antigens by Helicobacter pylori
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2011 (English)In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 83, no 16, p. 6336-6341Article in journal (Refereed) Published
Abstract [en]

Infectious diseases are often initiated by microbial adherence that is mediated by the binding of attachment molecules, termed adhesins, to cell surface receptors on host cells. We present an experimental system, oblique-incidence reflectivity difference (OI-RD) microscopy, which allows the detection of novel, low-affinity microbial attachment mechanisms that may be essential for infectious processes. OI-RD microscopy was used to analyze direct binding of the oncopathogen, Helicobacter pylori ( H. pylori ) to immobilized glycoconjugates in real time with no need for labeling tags. The results suggest the presence of additional Lewis b blood group antigen (Le(b)) binding adhesins that have not been detected previously. OI-RD microscopy also confirmed the high-affinity binding of H. pylori outer-membrane protein BabA to Le(b). The OI-RD microscopy method is broadly applicable to real-time characterization of intact microbial binding to host receptors and offers new strategies to elucidate the molecular interactions of infectious agents with human host cells.

Place, publisher, year, edition, pages
American Chemical Society (ACS), 2011
National Category
Chemical Sciences
Identifiers
urn:nbn:se:umu:diva-82538 (URN)10.1021/ac201260c (DOI)000293758800032 ()21721569 (PubMedID)
Funder
Swedish Research Council, 11218Swedish Cancer SocietyNIH (National Institute of Health), R01 AI070803, R01 AI081037, R01 HG003827-04, R01 GM076360-04S1
Available from: 2013-11-05 Created: 2013-11-05 Last updated: 2018-06-08Bibliographically approved
Lövgren, M., Bylund, G., Srivastava, M., Lundberg, C., Persson, O., Wingsle, G. & Wikström, M. (2004). The PRC-barrel domain of the ribosome maturation protein RimM mediates binding to ribosomal protein S19 in the 30S ribosomal subunits. RNA: A publication of the RNA Society, 10(11), 1798-1812
Open this publication in new window or tab >>The PRC-barrel domain of the ribosome maturation protein RimM mediates binding to ribosomal protein S19 in the 30S ribosomal subunits
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2004 (English)In: RNA: A publication of the RNA Society, ISSN 1355-8382, E-ISSN 1469-9001, Vol. 10, no 11, p. 1798-1812Article in journal (Refereed) Published
Abstract [en]

The RimM protein in Escherichia coli is associated with free 30S ribosomal subunits but not with 70S ribosomes. A DeltarimM mutant is defective in 30S maturation and accumulates 17S rRNA. To study the interaction of RimM with the 30S and its involvement in 30S maturation, RimM amino acid substitution mutants were constructed. A mutant RimM (RimM-YY-->AA), containing alanine substitutions for two adjacent tyrosines within the PRC beta-barrel domain, showed a reduced binding to 30S and an accumulation of 17S rRNA compared to wild-type RimM. The (RimM-YY-->AA) and DeltarimM mutants had significantly lower amounts of polysomes and also reduced levels of 30S relative to 50S compared to a wild-type strain. A mutation in rpsS, which encodes r-protein S19, suppressed the polysome- and 16S rRNA processing deficiencies of the RimM-YY-->AA but not that of the DeltarimM mutant. A mutation in rpsM, which encodes r-protein S13, suppressed the polysome deficiency of both rimM mutants. Suppressor mutations, found in either helices 31 or 33b of 16S rRNA, improved growth of both the RimM-YY-->AA and DeltarimM mutants. However, they suppressed the 16S rRNA processing deficiency of the RimM-YY-->AA mutant more efficiently than that of the DeltarimM mutant. Helices 31 and 33b are known to interact with S13 and S19, respectively, and S13 is known to interact with S19. A GST-RimM but not a GST-RimM(YY-->AA) protein bound strongly to S19 in 30S. Thus, RimM likely facilitates maturation of the region of the head of 30S that contains S13 and S19 as well as helices 31 and 33b.

Place, publisher, year, edition, pages
Cold Spring Harbor Laboratory Press (CSHL), 2004
Keywords
Alanine/metabolism, Amino Acid Sequence, Amino Acid Substitution, Bacterial Proteins/*chemistry/genetics/*metabolism, Escherichia coli/genetics/growth & development, Escherichia coli Proteins/*chemistry/genetics/*metabolism, Gene Expression Regulation; Bacterial, Glutathione Transferase/metabolism, Models; Molecular, Molecular Sequence Data, Mutagenesis; Site-Directed, Mutation, Protein Structure; Tertiary, RNA Processing; Post-Transcriptional, RNA; Ribosomal; 16S/genetics/metabolism, RNA-Binding Proteins, Recombinant Proteins/metabolism, Ribosomal Proteins/*chemistry/genetics/*metabolism, Ribosomes/*metabolism, Sequence Homology; Amino Acid, Tyrosine/metabolism
National Category
Medical and Health Sciences Forest Science Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-16783 (URN)10.1261/rna.7720204 (DOI)000224746300015 ()15496525 (PubMedID)
Available from: 2007-10-11 Created: 2007-10-11 Last updated: 2019-01-24Bibliographically approved
Bylund, G. O., Lövgren, J. M. & Wikström, P. M. (2001). Characterization of mutations in the metY-nusA-infB operon that suppress the slow growth of a DeltarimM mutant. Journal of Bacteriology, 183(20), 6095-6106
Open this publication in new window or tab >>Characterization of mutations in the metY-nusA-infB operon that suppress the slow growth of a DeltarimM mutant
2001 (English)In: Journal of Bacteriology, ISSN 0021-9193, E-ISSN 1098-5530, Vol. 183, no 20, p. 6095-6106Article in journal (Refereed) Published
Abstract [en]

The RimM protein in Escherichia coli is associated with free 30S ribosomal subunits but not with 70S ribosomes. A DeltarimM mutant shows a sevenfold-reduced growth rate and a reduced translational efficiency, probably as a result of aberrant assembly of the ribosomal 30S subunits. The slow growth and translational deficiency can be partially suppressed by increased synthesis of the ribosome binding factor RbfA. Here, we have identified 14 chromosomal suppressor mutations that increase the growth rate of a DeltarimM mutant by increasing the expression of rbfA. Nine of these mutations were in the nusA gene, which is located upstream from rbfA in the metY-nusA-infB operon; three mutations deleted the transcriptional terminator between infB and rbfA; one was an insertion of IS2 in infB, creating a new promoter for rbfA; and one was a duplication, placing a second copy of rbfA downstream from a promoter for the yhbM gene. Two of the nusA mutations were identical, while another mutation (nusA98) was identical to a previously isolated mutation, nusA11, shown to decrease termination of transcription. The different nusA mutations were found to increase the expression of rbfA by increasing the read-through of two internal transcriptional terminators located just downstream from the metY gene and that of the internal terminator preceding rbfA. Induced expression of the nusA(+) gene from a plasmid in a nusA(+) strain decreased the read-through of the two terminators just downstream from metY, demonstrating that one target for a previously proposed NusA-mediated feedback regulation of the metY-nusA-infB operon expression is these terminators. All of the nusA mutations produced temperature-sensitive phenotypes of rimM(+) strains. The nusA gene has previously been shown to be essential at 42 degrees C and below 32 degrees C. Here, we show that nusA is also essential at 37 degrees C.

Place, publisher, year, edition, pages
American Society for Microbiology, 2001
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:umu:diva-4315 (URN)10.1128/JB.183.20.6095-6106.2001 (DOI)000171267100035 ()11567010 (PubMedID)
Available from: 2004-12-15 Created: 2004-12-15 Last updated: 2019-01-21Bibliographically approved
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