263,704 research outputs found
Elucidation and control of substrate recognition during RiPP biosynthesis
No full textRibosomally synthesized and posttranslationally modified peptides (RiPPs) are a rapidly growing class of natural products. RiPP precursor peptides can undergo extensive enzymatic tailoring to yield structurally and functionally diverse products. Cyclodehydratases are a type of RiPP modifying enzyme that catalyze phosphorylation of the peptide backbone and subsequent nucleophilic attack by the sidechains of Cys, Ser, or Thr to form azoline heterocycless (or azoles upon oxidation). The catalytic unit of the cyclodehydratase is a YcaO-family protein which is often accompanied by a partner protein from the E1-like superfamily (depending on the of type cyclodehydratase). Although primarily known for azoline formation, recent work suggests that YcaO proteins can use different nucleophiles and partner proteins to generate thioamide, macroamidine, and possibly other peptide posttranslational modifications. In Chapter 1, I comprehensively review the biosynthetic gene clusters (BGCs), natural products, functions, mechanisms, and applications of YcaO proteins and outline future directions for this protein superfamily.
In Chapter 2, I report my investigations into the substrate recognition of canonical cyclodehydratases. Recent work suggested that unrelated RiPP modifying enzymes contain structurally similar precursor peptide binding domains. Using profile hidden Markov model comparisons, I discovered related and previously unrecognized peptide binding domains in proteins spanning the majority of known prokaryotic RiPP classes. This conserved domain was designated the RiPP precursor peptide recognition element (RRE). Through binding studies, I verified the RRE's role for three distinct RiPP classes: linear azole-containing peptides, thiopeptides, and lasso peptides. Because numerous RiPP biosynthetic enzymes act on peptide substrates, these findings have powerful predictive value as to which protein(s) drive substrate binding, thereby laying a foundation for further characterization of RiPP biosynthetic pathways and the rational engineering of new peptide binding activities.
In Chapter 3, I use knowledge gained from precursor peptide binding studies to engineer a peptide that can be recognized and modified by two biosynthetic enzymes from different pathways. Combining enzymes from multiple pathways is an attractive approach for producing molecules with desired structural features, but this strategy thus far has been hampered by limited substrate tolerance of enzymes from unrelated pathways. Because RiPP biosynthetic enzymes modify their substrates by binding motifs located usually in the N-terminal leader region of precursor peptides, RiPP biosynthetic systems are highly amenable to the engineering of new compounds. I exploit this by designing chimeric leader peptides that can be bound and processed by multiple enzymes from unrelated RiPP pathways. Using this broadly applicable strategy, a cyclodehydratase was combined with enzymes from the sactipeptide and lanthipeptide RiPP classes to create new-to-nature hybrid RiPPs. These hybrids provide insight into biosynthetic timing and enzyme compatibility and establish a general platform for the engineering additional hybrid RiPPs.Submission original under an indefinite embargo labeled 'Open Access'. The submission was exported from vireo on 2017-09-29 without embargo termsThe student, Brandon Burkhart, accepted the attached license on 2017-07-08 at 10:25.The student, Brandon Burkhart, submitted this Dissertation for approval on 2017-07-08 at 10:33.This Dissertation was approved for publication on 2017-07-10 at 11:37.DSpace SAF Submission Ingestion Package generated from Vireo submission #11344 on 2017-09-29 at 11:28:27Made available in DSpace on 2017-09-29T17:56:33Z (GMT). No. of bitstreams: 3
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Previous issue date: 2017-07-1
Genome-led discovery of novel RiPP natural products
No full textRibosomally synthesised and post-translationally modified peptides (RiPPs) are a structurally diverse class of natural product that display a range of clinically relevant bioactivities such as antimicrobial and anticancer. The full extent of RiPP biochemical complexity has not yet been fully explored. This is partly due to challenges associated with genome mining for RiPP biosynthetic gene clusters (BGCs), which is often hampered by poor detection of the short precursor peptides that are ultimately modified into the final metabolite. Microorganisms are therefore predicted to produce many more RiPPs than are currently known.
In this thesis, a novel RiPP genome mining tool, RiPPER, is employed to identify novel RiPP precursor peptides near YcaO-domain proteins, enzymes that catalyse various RiPP post-translational modifications including heterocyclisation and thioamidation. From this analysis, I report the identification of a novel and diverse family of RiPP BGCs present in over 230 species of Actinobacteria and Firmicutes.
A representative BGC from Streptomyces albus J1074 was characterised through cloning and expression of the pathway followed by genetic, metabolomic and structural studies. I thus report the identification of a novel RiPP, streptamidine, which contains a structurally rare amidine ring. The identification of this metabolite along with bioinformatic analysis of homologous pathways suggests that amidine-containing RiPPs might be widespread in nature, where previously only two examples have been characterised. Bottromycin and klebsazolicin are both antibiotic RiPPs whose activity has been attributed to their amidine rings. Amidines might therefore represent an important structural feature for antibiotics that can be explored in the future.
Overall, these studies show that many more biochemically diverse natural products can be discovered through the use of targeted genome mining approaches, even in widely studied model organisms such as Streptomyces albus J1074. This represents an exciting prospect for the future of antimicrobial discovery
RiPP Discovery and Regulation in Streptococcus thermophilus
No full textMounting bacterial resistance to antibiotics over the past two decades has imperiled their therapeutic efficacy. Since the majority of antibiotics currently used are derived from bacterial secondary metabolites, the discovery of new secondary metabolites may lead to the development of new, effective antibiotics. Ribosomally synthesized and post-translationally modified peptides (RiPPs) have recently emerged as a prominent class of secondary metabolites in bacteria, though RiPP diversity and regulation are not yet well understood. To these ends, I focus here on the str and kgr RiPP biosynthetic gene clusters (BGCs), which are both thought be regulated by specific short hydrophobic peptide (SHP) signaling, in the unstudied bacterium Streptococcus thermophilus JIM 8232. First, I show that S. thermophilus JIM 8232 produces the known secondary metabolite streptide and the cognate str SHP. It appears that the str SHP turns on streptide production towards mid-log growth phase. Second, targeted metabolomics suggests that the kgr BGC may not be expressed under normal growth conditions. Third, I report a tentative interaction between the kgr and str biosynthetic loci in S. thermophilus JIM 8232. Finally, I demonstrate progress towards identifying the RiPP that the kgr BGC encodes. Taken together, this work presents the first experimental characterization of RiPPs in S. thermophilus JIM 8232 and may provide a framework for leveraging SHP signaling for future secondary metabolite discovery in this strain
Matching of mass sequence tags to RiPP gene clusters using RiPP2Path.
No full text<p>Seven out of the nine search tags resulted in unique matches in their corresponding <i>Streptomyces</i> genomes.</p><p>Matching of mass sequence tags to RiPP gene clusters using RiPP2Path.</p
Expanding the structural diversity of the RiPP class of natural products
No full textRibosomally synthesized and post-translationally modified peptides (RiPPs) are a diverse and rapidly expanding class of natural products. Despite starting out as linear chains of amino acids, RiPP natural products acquire structural diversity ranging from small molecules like microcin C7 to 48-mer polytheonamides with varying post-translational modifications installed by biosynthetic enzymes. These conformational restrictions allow these natural products to exhibit a wide range of therapeutic activities like antifungal, antibacterial, allelopathic, and antiviral. Despite the structural and functional disparities, they all share a common feature: the presence of amino acid sequence in the precursor peptide called the 'leader peptide' that helps guide the biosynthetic enzymes towards the remaining precursor peptide termed the 'core peptide' to act on specific residues and install different PTMs. This leader-peptide guided biosynthetic route provides tremendous opportunity to bioengineer libraries of analogues of RiPPs. Exploiting this feature, in Chapter 2, I report my investigations on a strategy wherein by swapping the leader peptides from unrelated RiPP biosynthetic pathways and modifying the core peptide sequences, different biosynthetic enzymes can perform their characteristic chemistries on the core peptide residues.
In Chapter 3, I report the design and usage of chimeric leader peptides that can be bound by multiple enzymes from unrelated RiPP pathways to create new-to-nature hybrid RiPPs. This strategy involves the combination of a cyclodehydratase with modification enzymes from different lanthipeptide RiPP classes. These hybrids provide insight into enzyme promiscuity and compatibility, and the biosynthetic order of events, and establish a general platform for the engineering additional hybrid RiPPs.
In Chapter 4, I develop two different strategies for non-proteinogenic amino acid incorporation in the class I lantibiotic nisin and the class II lantibiotic lacticin 481 using amber stop codon technology.U of I OnlyAuthor requested U of Illinois access only (OA after 2yrs) in Vireo ETD syste
Expanding the structural diversity of the RiPP class of natural products
No full textRibosomally synthesized and post-translationally modified peptides (RiPPs) are a diverse and rapidly expanding class of natural products. Despite starting out as linear chains of amino acids, RiPP natural products acquire structural diversity ranging from small molecules like microcin C7 to 48-mer polytheonamides with varying post-translational modifications installed by biosynthetic enzymes. These conformational restrictions allow these natural products to exhibit a wide range of therapeutic activities like antifungal, antibacterial, allelopathic, and antiviral. Despite the structural and functional disparities, they all share a common feature: the presence of amino acid sequence in the precursor peptide called the 'leader peptide' that helps guide the biosynthetic enzymes towards the remaining precursor peptide termed the 'core peptide' to act on specific residues and install different PTMs. This leader-peptide guided biosynthetic route provides tremendous opportunity to bioengineer libraries of analogues of RiPPs. Exploiting this feature, in Chapter 2, I report my investigations on a strategy wherein by swapping the leader peptides from unrelated RiPP biosynthetic pathways and modifying the core peptide sequences, different biosynthetic enzymes can perform their characteristic chemistries on the core peptide residues.
In Chapter 3, I report the design and usage of chimeric leader peptides that can be bound by multiple enzymes from unrelated RiPP pathways to create new-to-nature hybrid RiPPs. This strategy involves the combination of a cyclodehydratase with modification enzymes from different lanthipeptide RiPP classes. These hybrids provide insight into enzyme promiscuity and compatibility, and the biosynthetic order of events, and establish a general platform for the engineering additional hybrid RiPPs.
In Chapter 4, I develop two different strategies for non-proteinogenic amino acid incorporation in the class I lantibiotic nisin and the class II lantibiotic lacticin 481 using amber stop codon technology.Submission published under a 24 month embargo labeled 'U of I Access', the embargo will last until 2020-05-01The student, Nidhi Kakkar, accepted the attached license on 2018-02-20 at 13:14.The student, Nidhi Kakkar, submitted this Dissertation for approval on 2018-02-20 at 13:25.This Dissertation was approved for publication on 2018-02-23 at 10:53.DSpace SAF Submission Ingestion Package generated from Vireo submission #12043 on 2018-08-31 at 17:17:08Made available in DSpace on 2018-09-04T20:33:48Z (GMT). No. of bitstreams: 2
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Previous issue date: 2018-02-23Embargo set by: Seth Robbins for item 107198
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Pueblos indígenas y capitalismo: ¿qué juego jugar y con qué reglas?
No full textEste monográfico de la Revista Internacional de Pensamiento Político (RIPP), dedicado a los pueblos indígenas surge de la mesa redonda “Derechos de los pueblos indígenas, lo común y la Naturaleza en contextos de pandemia”, que organizamos en el Salón de Grados de la Facultad de Derecho de la Universidad de Sevilla el día 4 de julio de 2019, dentro del I Congreso Derechos Humanos y Globalizació
Going Beyond Counting First Authors in Author Co-citation Analysis
No full textThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Research fronts in library and information science in Spain
No full textPublications and author cocitations in library and information science in Spain during the period from 1985 to 1994 were analyzed as a measure of the structure, specificity and composition of research fronts in this country. A cocitation matrix developed from an ad hoc database was subjected to cluster analysis, multidimensional scaling and principal components analysis, The resulting cocitation maps identified specific areas of r~search and their knowledge bases. We inferred the degree of consolidation of the discipline of library and information science, and of the subdisciplines informetrics, librarianship and university affiliation, from the research activities revealed. In this respect, the conclusions from the study show the existence of several research fronts in Spanish literature the contents of which are in most cases difficult to compare with those in other countries. A lesser degree of maturity of research in this field is shown
Protecting Animals 36: Author Witi Ihimaera
No full textIn this very special episode of Knowing Animals I am joined by beloved New Zealand author Witi Ihimaera. Witi has written many books featuring nonhuman animals. He offers us a non-colonial lens through which to think about the human/nonhuman relationship
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