Results
- Showing results for:
- Reset all filters
Search results
-
Journal articleCheng K, Wilkinson M, Chaban Y, et al., 2020,
A conformational switch in response to Chi converts RecBCD from phage destruction to DNA repair
, Nature Structural and Molecular Biology, Vol: 27, Pages: 71-77, ISSN: 1545-9985The RecBCD complex plays key roles in phage DNA degradation, CRISPR array acquisition (adaptation) and host DNA repair. The switch between these roles is regulated by a DNA sequence called Chi. We report cryo-EM structures of the Escherichia coli RecBCD complex bound to several different DNA forks containing a Chi sequence, including one in which Chi is recognized and others in which it is not. The Chi-recognized structure shows conformational changes in regions of the protein that contact Chi and reveals a tortuous path taken by the DNA. Sequence specificity arises from interactions with both the RecC subunit and the sequence itself. These structures provide molecular details for how Chi is recognized and insights into the changes that occur in response to Chi binding that switch RecBCD from bacteriophage destruction and CRISPR spacer acquisition to constructive host DNA repair.
-
Journal articleKopniczky MB, Canavan C, McClymont DW, et al., 2020,
Cell-free protein synthesis as a prototyping platform for mammalian synthetic biology
, ACS Synthetic Biology, Vol: 9, Pages: 144-156, ISSN: 2161-5063The field of mammalian synthetic biology is expanding quickly, and technologies for engineering large synthetic gene circuits are increasingly accessible. However, for mammalian cell engineering, traditional tissue culture methods are slow and cumbersome, and are not suited for high-throughput characterization measurements. Here we have utilized mammalian cell-free protein synthesis (CFPS) assays using HeLa cell extracts and liquid handling automation as an alternative to tissue culture and flow cytometry-based measurements. Our CFPS assays take a few hours, and we have established optimized protocols for small-volume reactions using automated acoustic liquid handling technology. As a proof-of-concept, we characterized diverse types of genetic regulation in CFPS, including T7 constitutive promoter variants, internal ribosomal entry sites (IRES) constitutive translation-initiation sequence variants, CRISPR/dCas9-mediated transcription repression, and L7Ae-mediated translation repression. Our data shows simple regulatory elements for use in mammalian cells can be quickly prototyped in a CFPS model system.
-
Journal articleSun Y, McCorvie TJ, Yates LA, et al., 2020,
Structural basis of homologous recombination
, CELLULAR AND MOLECULAR LIFE SCIENCES, Vol: 77, Pages: 3-18, ISSN: 1420-682X- Cite
- Citations: 61
-
Journal articlede Martín Garrido N, Crone MA, Ramlaul K, et al., 2020,
Bacteriophage MS2 displays unreported capsid variability assembling T = 4 and mixed capsids
, Molecular Microbiology, Vol: 113, Pages: 143-152, ISSN: 0950-382XBacteriophage MS2 is a positive-sense, single-stranded RNA virus encapsulated in an asymmetric T = 3 pseudo-icosahedral capsid. It infects Escherichia coli through the F-pilus, which it binds through a maturation protein incorporated into its capsid. Cryogenic electron microscopy has previously shown that its genome is highly ordered within virions, and that it regulates the assembly process of the capsid. In this study we have assembled recombinant MS2 capsids with non-genomic RNA containing the capsid incorporation sequence, and investigated the structures formed, revealing that T = 3, T = 4 and mixed capsids between these two triangulation numbers are generated, and resolving structures of T = 3 and T = 4 capsids to 4 Å and 6 Å respectively. We conclude that the basic MS2 capsid can form a mix of T = 3 and T = 4 structures, supporting a role for the ordered genome in favouring the formation of functional T = 3 virions.
-
Conference paperWebb AJ, Kelwick R, Wang Y, et al., 2019,
AL-PHA beads: bioplastic-bsaed protease biosensors for global health
, British Society for Parasitology Autumn Symposium, Belfast, UK -
Journal articleLai H-E, Canavan C, Cameron L, et al., 2019,
Synthetic biology and the United Nations
, Trends in Biotechnology, Vol: 37, Pages: 1146-1151, ISSN: 0167-7799Synthetic biology is a rapidly emerging interdisciplinary field of science and engineering that aims to redesign living systems through reprogramming genetic information. The field has catalysed global debate among policymakers and publics. Here we describe how synthetic biology relates to these international deliberations, particularly the Convention on Biological Diversity (CBD).
-
Journal articleYe F, Kotta-Loizou I, Jovanovic M, et al., 2019,
Structural basis of transcription inhibition by the DNA mimic protein Ocr of bacteriophage T7
, eLife, Vol: 9, ISSN: 2050-084XAbstract Bacteriophage T7 infects Escherichia coli and evades the host defence system. The Ocr protein of T7 was shown to exist as a dimer mimicking DNA and to bind to host restriction enzymes, thus preventing the degradation of the viral genome by the host. Here we report that Ocr can also inhibit host transcription by directly binding to bacterial RNA polymerase (RNAP) and competing with the recruitment of RNAP by sigma factors. Using cryo electron microscopy, we determined the structures of Ocr bound to RNAP. The structures show that an Ocr dimer binds to RNAP in the cleft, where key regions of sigma bind and where DNA resides during transcription synthesis, thus providing a structural basis for the transcription inhibition. Our results reveal the versatility of Ocr in interfering with host systems and suggest possible strategies that could be exploited in adopting DNA mimicry as a basis for forming novel antibiotics. Impact statement DNA mimicry Ocr protein, a well-studied T7 phage protein that inhibits host restriction enzymes, can also inhibit host transcription through competing with sigma factors in binding to RNA polymerase.
-
Journal articleRiglar DT, Richmond DL, Potvin-Trottier L, et al., 2019,
Bacterial variability in the mammalian gut captured by a single-cell synthetic oscillator
, NATURE COMMUNICATIONS, Vol: 10- Author Web Link
- Cite
- Citations: 39
-
Journal articleFreemont P, 2019,
Synthetic biology industry - Data-driven design is creating new opportunities in biotechnology.
, Emerging Topics in Life Sciences, Vol: 3, Pages: 651-657, ISSN: 2397-8554Synthetic biology is a rapidly emerging interdisciplinary research field that is primarily built upon foundational advances in molecular biology combined with engineering design. The field considers living systems as programmable at the genetic level and has been defined by the development of new platform technologies. This has spurned a rapid growth in start-up companies and the new synthetic biology industry is growing rapidly, with start-up companies receiving ~$6.1B investment since 2015 and a global synthetic biology market value estimated to be $14B by 2026. Many of the new start-upscan be grouped within a multi-layer ‘technology stack’. The ‘stack’ comprises a number of technology layers which together can be applied to a diversity of new biotechnology applications like consumer biotechnology products and living therapies. The ‘stack’ also enables new commercial opportunities and value chains similar to the software design and manufacturing revolution of the 20th century. However, synthetic biology industry is at a crucial point, as it now requires recognisable commercial successes in order for the industry to expand and scale, in terms of investment and companies. However, such expansion may directly challenge the ethos of synthetic biology, in terms of open technology sharing and democratisation, which could by accident lead to multi-national corporations and technology monopolies similar to the existing biotechnology/biopharma industry.
-
Journal articleWood TE, Howard SA, Forster A, et al., 2019,
The Pseudomonas aeruginosa T6SS delivers a periplasmic toxin that disrupts bacterial cell morphology
, Cell Reports, Vol: 29, Pages: 187-201.e7, ISSN: 2211-1247The type VI secretion system (T6SS) is crucialin interbacterial competition and is avirulence determinant ofmany Gram-negative bacteria. Several T6SS effectorsarecovalently fused to secreted T6SS structural components such asthe VgrG spike for delivery into target cells.In Pseudomonas aeruginosa, theVgrG2b effector waspreviously proposedto mediatebacterial internalisation into eukaryotic cells. In this work, wefind that the VgrG2b C-terminal domain(VgrG2bC-ter) elicits toxicity in the bacterial periplasm, counteracted by a cognate immunity protein.We resolve thestructure of VgrG2bC-ter and confirm it is a member ofthezinc-metallopeptidasefamily of enzymes. We show that this effector causesmembrane blebbing atmidcell, whichsuggests a distincttype of T6SS-mediated growthinhibition through interference with cell division, mimicking the impact of β-lactam antibiotics. Ourstudyintroduces a further effector family to the T6SS arsenaland demonstrates that VgrG2b can target both prokaryotic and eukaryotic cells.
This data is extracted from the Web of Science and reproduced under a licence from Thomson Reuters. You may not copy or re-distribute this data in whole or in part without the written consent of the Science business of Thomson Reuters.
General enquiries
Section Manager
Brett Onslow
+44 (0)20 7594 3871
Personal Assistant for the Section of Structural Biology
Kasia Pearce
+44 (0)20 7594 2917
Laboratory Manager
Soo Mei Chee