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  1. Article ; Online: Bioinformatic characterization of endolysins and holin-like membrane proteins in the lysis cassette of phages that infect Gordonia rubripertincta.

    Richard S Pollenz / Jackson Bland / Welkin H Pope

    PLoS ONE, Vol 17, Iss 11, p e

    2022  Volume 0276603

    Abstract: Holins are bacteriophage-encoded transmembrane proteins that function to control the timing of bacterial lysis event, assist with the destabilization of the membrane proton motive force and in some models, generate large "pores" in the cell membrane to ... ...

    Abstract Holins are bacteriophage-encoded transmembrane proteins that function to control the timing of bacterial lysis event, assist with the destabilization of the membrane proton motive force and in some models, generate large "pores" in the cell membrane to allow the exit of the phage-encoded endolysin so they can access the peptidoglycan components of the cell wall. The lysis mechanism has been rigorously evaluated through biochemical and genetic studies in very few phages, and the results indicate that phages utilize endolysins, holins and accessory proteins to the outer membrane to achieve cell lysis through several distinct operational models. This observation suggests the possibility that phages may evolve novel variations of how the lysis proteins functionally interact in an effort to improve fitness or evade host defenses. To begin to address this hypothesis, the current study utilized a comprehensive bioinformatic approach to systematically identify the proteins encoded by the genes within the lysis cassettes in 16 genetically diverse phages that infect the Gram-positive Gordonia rubripertincta NRLL B-16540 strain. The results show that there is a high level of diversity of the various lysis genes and 16 different genome organizations of the putative lysis cassette, many which have never been described. Thirty-four different genes encoding holin-like proteins were identified as well as a potential holin-major capsid fusion protein. The holin-like proteins contained between 1-4 transmembrane helices, were not shared to a high degree amongst the different phages and are present in the lysis cassette in a wide range of combinations of up to 4 genes in which none are duplicated. Detailed evaluation of the transmembrane domains and predicted membrane topologies of the holin-like proteins show that many have novel structures that have not been previously characterized. These results provide compelling support that there are novel operational lysis models yet to be discovered.
    Keywords Medicine ; R ; Science ; Q
    Subject code 612
    Language English
    Publishing date 2022-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  2. Article ; Online: Mass Spectrometry as a Tool to Enhance "-omics" Education

    Michael J. Wolyniak / Nathan S. Reyna / Ruth Plymale / Welkin H. Pope / Daniel E. Westholm

    Journal of Microbiology & Biology Education, Vol 19, Iss

    2018  Volume 1

    Abstract: The rise of "-omics" related technologies makes it essential for students to have experience working with large bioinformatics data sets. Although”-omic” datasets are complex and abstract, effective instruction can be improved when students see the ... ...

    Abstract The rise of "-omics" related technologies makes it essential for students to have experience working with large bioinformatics data sets. Although”-omic” datasets are complex and abstract, effective instruction can be improved when students see the direct connections between the data on a computer screen and the results of "wet lab" experimentation. Here we describe the use of protein mass spectrometry as a means for students to gain experience in connecting bioinformatic data with work done at the lab bench. Course-based Research Experiences (CREs) based on these techniques are accessible to institutions of all types as a result of rapidly declining costs for whole genome and proteome analysis. Our implementation is within a CRE based on viral infection of a bacterial host; however, this basic paradigm may be applied to other experimental systems of interest.
    Keywords mass spectrometry ; -omics ; active learning ; laboratory skills ; Special aspects of education ; LC8-6691 ; Biology (General) ; QH301-705.5
    Language English
    Publishing date 2018-05-01T00:00:00Z
    Publisher American Society for Microbiology
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  3. Article ; Online: Mass Spectrometry as a Tool to Enhance “-omics” Education

    Michael J. Wolyniak / Nathan S. Reyna / Ruth Plymale / Welkin H. Pope / Daniel E. Westholm

    Journal of Microbiology & Biology Education, Vol 19, Iss

    2018  Volume 1

    Abstract: The rise of "-omics" related technologies makes it essential for students to have experience working with large bioinformatics data sets. Although”-omic” datasets are complex and abstract, effective instruction can be improved when students see the ... ...

    Abstract The rise of "-omics" related technologies makes it essential for students to have experience working with large bioinformatics data sets. Although”-omic” datasets are complex and abstract, effective instruction can be improved when students see the direct connections between the data on a computer screen and the results of "wet lab" experimentation. Here we describe the use of protein mass spectrometry as a means for students to gain experience in connecting bioinformatic data with work done at the lab bench. Course-based Research Experiences (CREs) based on these techniques are accessible to institutions of all types as a result of rapidly declining costs for whole genome and proteome analysis. Our implementation is within a CRE based on viral infection of a bacterial host; however, this basic paradigm may be applied to other experimental systems of interest.
    Keywords Special aspects of education ; LC8-6691 ; Biology (General) ; QH301-705.5
    Language English
    Publishing date 2018-03-01T00:00:00Z
    Publisher American Society for Microbiology
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  4. Article ; Online: Whole genome comparison of a large collection of mycobacteriophages reveals a continuum of phage genetic diversity

    Welkin H Pope / Charles A Bowman / Daniel A Russell / Deborah Jacobs-Sera / David J Asai / Steven G Cresawn / William R Jacobs Jr / Roger W Hendrix / Jeffrey G Lawrence / Graham F Hatfull / Science Education Alliance Phage Hunters Advancing Genomics and Evolutionary Science / Phage Hunters Integrating Research and Education / Mycobacterial Genetics Course

    eLife, Vol

    2015  Volume 4

    Abstract: The bacteriophage population is large, dynamic, ancient, and genetically diverse. Limited genomic information shows that phage genomes are mosaic, and the genetic architecture of phage populations remains ill-defined. To understand the population ... ...

    Abstract The bacteriophage population is large, dynamic, ancient, and genetically diverse. Limited genomic information shows that phage genomes are mosaic, and the genetic architecture of phage populations remains ill-defined. To understand the population structure of phages infecting a single host strain, we isolated, sequenced, and compared 627 phages of Mycobacterium smegmatis. Their genetic diversity is considerable, and there are 28 distinct genomic types (clusters) with related nucleotide sequences. However, amino acid sequence comparisons show pervasive genomic mosaicism, and quantification of inter-cluster and intra-cluster relatedness reveals a continuum of genetic diversity, albeit with uneven representation of different phages. Furthermore, rarefaction analysis shows that the mycobacteriophage population is not closed, and there is a constant influx of genes from other sources. Phage isolation and analysis was performed by a large consortium of academic institutions, illustrating the substantial benefits of a disseminated, structured program involving large numbers of freshman undergraduates in scientific discovery.
    Keywords bacteriophage ; genomic ; evolution ; Medicine ; R ; Science ; Q ; Biology (General) ; QH301-705.5
    Subject code 612
    Language English
    Publishing date 2015-04-01T00:00:00Z
    Publisher eLife Sciences Publications Ltd
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  5. Article ; Online: Genomic diversity of bacteriophages infecting Microbacterium spp.

    Deborah Jacobs-Sera / Lawrence A Abad / Richard M Alvey / Kirk R Anders / Haley G Aull / Suparna S Bhalla / Lawrence S Blumer / David W Bollivar / J Alfred Bonilla / Kristen A Butela / Roy J Coomans / Steven G Cresawn / Tom D'Elia / Arturo Diaz / Ashley M Divens / Nicholas P Edgington / Gregory D Frederick / Maria D Gainey / Rebecca A Garlena /
    Kenneth W Grant / Susan M R Gurney / Heather L Hendrickson / Lee E Hughes / Margaret A Kenna / Karen K Klyczek / Hari Kotturi / Travis N Mavrich / Angela L McKinney / Evan C Merkhofer / Jordan Moberg Parker / Sally D Molloy / Denise L Monti / Dana A Pape-Zambito / Richard S Pollenz / Welkin H Pope / Nathan S Reyna / Claire A Rinehart / Daniel A Russell / Christopher D Shaffer / Viknesh Sivanathan / Ty H Stoner / Joseph Stukey / C Nicole Sunnen / Sara S Tolsma / Philippos K Tsourkas / Jamie R Wallen / Vassie C Ware / Marcie H Warner / Jacqueline M Washington / Kristi M Westover

    PLoS ONE, Vol 15, Iss 6, p e

    2020  Volume 0234636

    Abstract: The bacteriophage population is vast, dynamic, old, and genetically diverse. The genomics of phages that infect bacterial hosts in the phylum Actinobacteria show them to not only be diverse but also pervasively mosaic, and replete with genes of unknown ... ...

    Abstract The bacteriophage population is vast, dynamic, old, and genetically diverse. The genomics of phages that infect bacterial hosts in the phylum Actinobacteria show them to not only be diverse but also pervasively mosaic, and replete with genes of unknown function. To further explore this broad group of bacteriophages, we describe here the isolation and genomic characterization of 116 phages that infect Microbacterium spp. Most of the phages are lytic, and can be grouped into twelve clusters according to their overall relatedness; seven of the phages are singletons with no close relatives. Genome sizes vary from 17.3 kbp to 97.7 kbp, and their G+C% content ranges from 51.4% to 71.4%, compared to ~67% for their Microbacterium hosts. The phages were isolated on five different Microbacterium species, but typically do not efficiently infect strains beyond the one on which they were isolated. These Microbacterium phages contain many novel features, including very large viral genes (13.5 kbp) and unusual fusions of structural proteins, including a fusion of VIP2 toxin and a MuF-like protein into a single gene. These phages and their genetic components such as integration systems, recombineering tools, and phage-mediated delivery systems, will be useful resources for advancing Microbacterium genetics.
    Keywords Medicine ; R ; Science ; Q
    Language English
    Publishing date 2020-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  6. Article ; Online: Cluster J mycobacteriophages

    Welkin H Pope / Deborah Jacobs-Sera / Aaron A Best / Gregory W Broussard / Pamela L Connerly / Rebekah M Dedrick / Timothy A Kremer / Susan Offner / Amenawon H Ogiefo / Marie C Pizzorno / Kate Rockenbach / Daniel A Russell / Emily L Stowe / Joseph Stukey / Sarah A Thibault / James F Conway / Roger W Hendrix / Graham F Hatfull

    PLoS ONE, Vol 8, Iss 7, p e

    intron splicing in capsid and tail genes.

    2013  Volume 69273

    Abstract: Bacteriophages isolated on Mycobacterium smegmatis mc(2)155 represent many distinct genomes sharing little or no DNA sequence similarity. The genomes are architecturally mosaic and are replete with genes of unknown function. A new group of genomes ... ...

    Abstract Bacteriophages isolated on Mycobacterium smegmatis mc(2)155 represent many distinct genomes sharing little or no DNA sequence similarity. The genomes are architecturally mosaic and are replete with genes of unknown function. A new group of genomes sharing substantial nucleotide sequences constitute Cluster J. The six mycobacteriophages forming Cluster J are morphologically members of the Siphoviridae, but have unusually long genomes ranging from 106.3 to 117 kbp. Reconstruction of the capsid by cryo-electron microscopy of mycobacteriophage BAKA reveals an icosahedral structure with a triangulation number of 13. All six phages are temperate and homoimmune, and prophage establishment involves integration into a tRNA-Leu gene not previously identified as a mycobacterial attB site for phage integration. The Cluster J genomes provide two examples of intron splicing within the virion structural genes, one in a major capsid subunit gene, and one in a tail gene. These genomes also contain numerous free-standing HNH homing endonuclease, and comparative analysis reveals how these could contribute to genome mosaicism. The unusual Cluster J genomes provide new insights into phage genome architecture, gene function, capsid structure, gene mobility, intron splicing, and evolution.
    Keywords Medicine ; R ; Science ; Q
    Subject code 572
    Language English
    Publishing date 2013-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  7. Article ; Online: Comparative genomics of Cluster O mycobacteriophages.

    Steven G Cresawn / Welkin H Pope / Deborah Jacobs-Sera / Charles A Bowman / Daniel A Russell / Rebekah M Dedrick / Tamarah Adair / Kirk R Anders / Sarah Ball / David Bollivar / Caroline Breitenberger / Sandra H Burnett / Kristen Butela / Deanna Byrnes / Sarah Carzo / Kathleen A Cornely / Trevor Cross / Richard L Daniels / David Dunbar /
    Ann M Findley / Chris R Gissendanner / Urszula P Golebiewska / Grant A Hartzog / J Robert Hatherill / Lee E Hughes / Chernoh S Jalloh / Carla De Los Santos / Kevin Ekanem / Sphindile L Khambule / Rodney A King / Christina King-Smith / Karen Klyczek / Greg P Krukonis / Christian Laing / Jonathan S Lapin / A Javier Lopez / Sipho M Mkhwanazi / Sally D Molloy / Deborah Moran / Vanisha Munsamy / Eddie Pacey / Ruth Plymale / Marianne Poxleitner / Nathan Reyna / Joel F Schildbach / Joseph Stukey / Sarah E Taylor / Vassie C Ware / Amanda L Wellmann / Daniel Westholm

    PLoS ONE, Vol 10, Iss 3, p e

    2015  Volume 0118725

    Abstract: Mycobacteriophages--viruses of mycobacterial hosts--are genetically diverse but morphologically are all classified in the Caudovirales with double-stranded DNA and tails. We describe here a group of five closely related mycobacteriophages--Corndog, ... ...

    Abstract Mycobacteriophages--viruses of mycobacterial hosts--are genetically diverse but morphologically are all classified in the Caudovirales with double-stranded DNA and tails. We describe here a group of five closely related mycobacteriophages--Corndog, Catdawg, Dylan, Firecracker, and YungJamal--designated as Cluster O with long flexible tails but with unusual prolate capsids. Proteomic analysis of phage Corndog particles, Catdawg particles, and Corndog-infected cells confirms expression of half of the predicted gene products and indicates a non-canonical mechanism for translation of the Corndog tape measure protein. Bioinformatic analysis identifies 8-9 strongly predicted SigA promoters and all five Cluster O genomes contain more than 30 copies of a 17 bp repeat sequence with dyad symmetry located throughout the genomes. Comparison of the Cluster O phages provides insights into phage genome evolution including the processes of gene flux by horizontal genetic exchange.
    Keywords Medicine ; R ; Science ; Q
    Language English
    Publishing date 2015-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  8. Article ; Online: Cluster K mycobacteriophages

    Welkin H Pope / Christina M Ferreira / Deborah Jacobs-Sera / Robert C Benjamin / Ariangela J Davis / Randall J DeJong / Sarah C R Elgin / Forrest R Guilfoile / Mark H Forsyth / Alexander D Harris / Samuel E Harvey / Lee E Hughes / Peter M Hynes / Arrykka S Jackson / Marilyn D Jalal / Elizabeth A MacMurray / Coreen M Manley / Molly J McDonough / Jordan L Mosier /
    Larissa J Osterbann / Hannah S Rabinowitz / Corwin N Rhyan / Daniel A Russell / Margaret S Saha / Christopher D Shaffer / Stephanie E Simon / Erika F Sims / Isabel G Tovar / Emilie G Weisser / John T Wertz / Kathleen A Weston-Hafer / Kurt E Williamson / Bo Zhang / Steven G Cresawn / Paras Jain / Mariana Piuri / William R Jacobs / Roger W Hendrix / Graham F Hatfull

    PLoS ONE, Vol 6, Iss 10, p e

    insights into the evolutionary origins of mycobacteriophage TM4.

    2011  Volume 26750

    Abstract: Five newly isolated mycobacteriophages--Angelica, CrimD, Adephagia, Anaya, and Pixie--have similar genomic architectures to mycobacteriophage TM4, a previously characterized phage that is widely used in mycobacterial genetics. The nucleotide sequence ... ...

    Abstract Five newly isolated mycobacteriophages--Angelica, CrimD, Adephagia, Anaya, and Pixie--have similar genomic architectures to mycobacteriophage TM4, a previously characterized phage that is widely used in mycobacterial genetics. The nucleotide sequence similarities warrant grouping these into Cluster K, with subdivision into three subclusters: K1, K2, and K3. Although the overall genome architectures of these phages are similar, TM4 appears to have lost at least two segments of its genome, a central region containing the integration apparatus, and a segment at the right end. This suggests that TM4 is a recent derivative of a temperate parent, resolving a long-standing conundrum about its biology, in that it was reportedly recovered from a lysogenic strain of Mycobacterium avium, but it is not capable of forming lysogens in any mycobacterial host. Like TM4, all of the Cluster K phages infect both fast- and slow-growing mycobacteria, and all of them--with the exception of TM4--form stable lysogens in both Mycobacterium smegmatis and Mycobacterium tuberculosis; immunity assays show that all five of these phages share the same immune specificity. TM4 infects these lysogens suggesting that it was either derived from a heteroimmune temperate parent or that it has acquired a virulent phenotype. We have also characterized a widely-used conditionally replicating derivative of TM4 and identified mutations conferring the temperature-sensitive phenotype. All of the Cluster K phages contain a series of well conserved 13 bp repeats associated with the translation initiation sites of a subset of the genes; approximately one half of these contain an additional sequence feature composed of imperfectly conserved 17 bp inverted repeats separated by a variable spacer. The K1 phages integrate into the host tmRNA and the Cluster K phages represent potential new tools for the genetics of M. tuberculosis and related species.
    Keywords Medicine ; R ; Science ; Q
    Subject code 572
    Language English
    Publishing date 2011-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  9. Article ; Online: Expanding the diversity of mycobacteriophages

    Welkin H Pope / Deborah Jacobs-Sera / Daniel A Russell / Craig L Peebles / Zein Al-Atrache / Turi A Alcoser / Lisa M Alexander / Matthew B Alfano / Samantha T Alford / Nichols E Amy / Marie D Anderson / Alexander G Anderson / Andrew A S Ang / Manuel Ares / Amanda J Barber / Lucia P Barker / Jonathan M Barrett / William D Barshop / Cynthia M Bauerle /
    Ian M Bayles / Katherine L Belfield / Aaron A Best / Agustin Borjon / Charles A Bowman / Christine A Boyer / Kevin W Bradley / Victoria A Bradley / Lauren N Broadway / Keshav Budwal / Kayla N Busby / Ian W Campbell / Anne M Campbell / Alyssa Carey / Steven M Caruso / Rebekah D Chew / Chelsea L Cockburn / Lianne B Cohen / Jeffrey M Corajod / Steven G Cresawn / Kimberly R Davis / Lisa Deng / Dee R Denver / Breyon R Dixon / Sahrish Ekram / Sarah C R Elgin / Angela E Engelsen / Belle E V English / Marcella L Erb / Crystal Estrada / Laura Z Filliger

    PLoS ONE, Vol 6, Iss 1, p e

    insights into genome architecture and evolution.

    2011  Volume 16329

    Abstract: Mycobacteriophages are viruses that infect mycobacterial hosts such as Mycobacterium smegmatis and Mycobacterium tuberculosis. All mycobacteriophages characterized to date are dsDNA tailed phages, and have either siphoviral or myoviral morphotypes. ... ...

    Abstract Mycobacteriophages are viruses that infect mycobacterial hosts such as Mycobacterium smegmatis and Mycobacterium tuberculosis. All mycobacteriophages characterized to date are dsDNA tailed phages, and have either siphoviral or myoviral morphotypes. However, their genetic diversity is considerable, and although sixty-two genomes have been sequenced and comparatively analyzed, these likely represent only a small portion of the diversity of the mycobacteriophage population at large. Here we report the isolation, sequencing and comparative genomic analysis of 18 new mycobacteriophages isolated from geographically distinct locations within the United States. Although no clear correlation between location and genome type can be discerned, these genomes expand our knowledge of mycobacteriophage diversity and enhance our understanding of the roles of mobile elements in viral evolution. Expansion of the number of mycobacteriophages grouped within Cluster A provides insights into the basis of immune specificity in these temperate phages, and we also describe a novel example of apparent immunity theft. The isolation and genomic analysis of bacteriophages by freshman college students provides an example of an authentic research experience for novice scientists.
    Keywords Medicine ; R ; Science ; Q
    Language English
    Publishing date 2011-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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