Characterisation of multiple substrate-specific (d)ITP/(d)XTPase and modelling of deaminated purine nucleotide metabolism

To be viable, organisms possess a number of (deoxy)nucleotide phosphohydrolases, which hydrolyze these nucleotides removing them from the active NTP and dNTP pools. Deamination of purine bases can result in accumulation of such nucleotides as ITP, dITP, XTP and dXTP. E. coli RdgB has been characterised as a deoxyribonucleoside triphosphate pyrophosphohydrolase that can act on these nucleotides. S. cerevisiae homologue encoded by YJR069C was purified and its (d)NTPase activity was assayed using fifteen nucleotide substrates. ITP, dITP, and XTP were identified as major substrates and kinetic parameters measured. Inhibition by ATP, dATP and GTP were established. On the basis of experimental and published data, modelling and simulation of ITP, dITP, XTP and dXTP metabolism was performed. (d)ITP/(d)XTPase is a new example of enzyme with multiple substrate-specificity demonstrating that multispecificity is not a rare phenomenon

Close-range photogrammetry enables documentation of environment-induced deformation of architectural heritage

Deformation, damage and permanent loss of heritage assets due to various physical and environmental factors has always been a major problem. As the availability of funds for conservation and restoration is limited, the digital documentation of heritage objects and monitoring of environment-induced deformations are increasingly important for cultural heritage preservation. Our study elucidates developments in the digital image capturing and processing for recording architectural heritage objects focusing on the digital camera calibration, close-range imaging, and photogrammetric modelling of complex structures using image matching techniques. A particular consideration in this paper is given to the ortho-photographic image compiling and accuracy assessment procedure. The practicality of the methodology is demonstrated by applying photogrammetric system PhotoMod for documentation of decorative elements in Uzutrakis manor, a national heritage site in Trakai, Lithuania

Close-range photogrammetry enables documentation of environment-induced deformation of architectural heritage

Deformation, damage and permanent loss of heritage assets due to various physical and environmental factors has always been a major problem. As the availability of funds for conservation and restoration is limited, the digital documentation of heritage objects and monitoring of environment-induced deformations are increasingly important for cultural heritage preservation. Our study elucidates developments in the digital image capturing and processing for recording architectural heritage objects focusing on the digital camera calibration, close-range imaging, and photogrammetric modelling of complex structures using image matching techniques. A particular consideration in this paper is given to the ortho-photographic image compiling and accuracy assessment procedure. The practicality of the methodology is demonstrated by applying photogrammetric system PhotoMod for documentation of decorative elements in Uzutrakis manor, a national heritage site in Trakai, Lithuania

Improving carbon monoxide tolerance of Cupriavidus necator H16 through adaptive laboratory evolution

Background: The toxic gas carbon monoxide (CO) is abundantly present in synthesis gas (syngas) and certain industrial waste gases that can serve as feedstocks for the biological production of industrially significant chemicals and fuels. For efficient bacterial growth to occur, and to increase productivity and titres, a high resistance to the gas is required. The aerobic bacterium Cupriavidus necator H16 can grow on CO2 + H2, although it cannot utilise CO as a source of carbon and energy. This study aimed to increase its CO resistance through adaptive laboratory evolution.Results: To increase the tolerance of C. necator to CO, the organism was continually subcultured in the presence of CO both heterotrophically and autotrophically. Ten individual cultures were evolved heterotrophically with fructose in this manner and eventually displayed a clear growth advantage over the wild type strain. Next-generation sequencing revealed several mutations, including a single point mutation upstream of a cytochrome bd ubiquinol oxidase operon (cydA2B2), which was present in all evolved isolates. When a subset of these mutations was engineered into the parental H16 strain, only the cydA2B2 upstream mutation enabled faster growth in the presence of CO. Expression analysis, mutation, overexpression and complementation suggested that cydA2B2 transcription is upregulated in the evolved isolates, resulting in increased CO tolerance under heterotrophic but not autotrophic conditions. However, through subculturing on a syngas-like mixture with increasing CO concentrations, C. necator could also be evolved to tolerate high CO concentrations under autotrophic conditions. A mutation in the gene for the soluble [NiFe]-hydrogenase subunit hoxH was identified in the evolved isolates. When the resulting amino acid change was engineered into the parental strain, autotrophic CO resistance was conferred. A strain constitutively expressing cydA2B2 and the mutated hoxH gene exhibited high CO tolerance under both heterotrophic and autotrophic conditions.Conclusion:C. necator was evolved to tolerate high concentrations of CO, a phenomenon which was dependent on the terminal respiratory cytochrome bd ubiquinol oxidase when grown heterotrophically and the soluble [NiFe]-hydrogenase when grown autotrophically. A strain exhibiting high tolerance under both conditions was created and presents a promising chassis for syngas-based bioproduction processes

Urban regeneration for sustainable communities: A case study

Over the past few decades, cities have been subjected to severe social and economic pressures, which have had a disproportionate impact on the urban environment. Consequently, this has increased the concentration of the most deprived households in the worst urban neighbourhoods. The UK Governments have attempted to tackle the physical, social and economic consequences of these changes through a variety of mechanisms and policy initiatives with varying degrees of success. The current objective can be interpreted as an attempt to implement regeneration projects or create sustainable communities/places through multi‐agency or partnership programmes. The nature and extent of urban problems are now very well documented; there has been a considerable learning process in the development of different urban regeneration programmes. Over the 20th and the beginning of 21st centuries, the nature of urban regeneration policy has changed direction several times and has applied different focuses. Today, a “sustainable community” is a key issue in an ambitious Government programme “umbrella”. As an objective of the present programme, the UK Government introduced the Sustainable Community Plan in 2003, describing a vision of how our communities are to be developed over next 20 years economically, socially and environmentally, while respecting the needs of future generations. The paper assesses how closely the Regeneration Programme is aligned to the Sustainable Communities Plan. The research within several surveys focuses on sustainable communities and urban regeneration, both have certain parallels and functional overlaps. Santrauka Šiuo metu siekiama įdiegti atgaivinimo projektus ir sukurti darniąsias bendruomenes pasitelkiant partnerystės programas. Darniosios bendruomenės kūrimas yra viena svarbiausių problemų, analizuojamų JK vyriausybės programoje. 2003 m. JK vyriausybes sukurtame darniųjų bendruomenių plane pateikiama vizija, kaip mūsų bendruomenės turetų vystytis ekonomiškai, socialiai ir aplinkos atžvilgiu per ateinančius 20 metu, atsižvelgiant į ateities kartų gerovę ir poreikius. Analizuojama, kaip glaustai miestų atgaivinimo programa yra suderinta su darniųjų bendruomenių kūrimo planu. Tyrimai, pagrįsti tiesioginėmis apklausomis, įvertina darniųjų bendruomenių ir miestų atgaivinimo pletrą bei sąsajas. First published online: 21 Oct 2010 Reikšminiai žodžiai: darnumas, bendruomenė, atgaivinimas, atvejo tyrimas, Castlefields, J

Commercial leisure property valuation: A comparison of the case studies in UK and Lithuania

The development of leisure as an economic activity has come into focus over the last century. Nowadays, a commercial leisure property has gained an important role within the total spectrum of world economies. Despite these developments in the leisure sector, the valuation of property for the purpose of commercial leisure is still regarded as an Achilles’ heel. The accepted approach to the property valuation in UK is to perform an open market valuation for the existing use. However, for the specialized properties including leisure assets, the distinctive valuation methodology has to be applied. This paper outlines the principles of methodology that are commonly used for the valuation of leisure property in the UK, Lithuania and other countries. Factors affecting the valuation process with respect to the property development and investment decision are discussed. The study is accompanied with examples of valuation method applications and analyses of individual case studies of golf course valuation. Santruka Per pastaraji šimtmeti laisvalaikio vystymas tapo reikšminga ekonomikos veiklos sritis. Šiuo metu laisvalaikio verslo nekilnojamasis turtas vaidina svarbu vaidmeni pasaulio šaliu ūkineje veikloje. Nepaisant laisvalaikio sektoriaus išsivystymo, nekilnojamojo turto vertinimas laisvalaikio verslo srityje vis dar yra silpnas. JK priimtas nekilnojamojo turto vertinimo būdas pagristas atviros rinkos vertinimu pagal turto paskirti. Tačiau specializuotam nekilnojamajam turtui, taip pat ir laisvalaikio, išskirtinis vertinimo metodas turi būti taikomas. Šiame straipsnyje apibūdinami principai, kurie taikomi laisvalaikio nekilnojamojo turto vertinimui JK, Lietuvoje ir kitose šalyse. Straipsnyje nagrinejami veiksniai, kurie daro itaka vertinimo procesui, kai nekilnojamasis turtas vertinamas vystymo ar investavimo tikslais. Tyrime pateikti golfo aikštynu vertinimo metodu ir analizes pavyzdžiai. First Published Online: 18 Oct 201

Translation initiation events on structured eukaryotic mRNAs generate gene expression noise

Gene expression stochasticity plays a major role in biology, creating non-genetic cellular individuality and influencing multiple processes, including differentiation and stress responses. We have addressed the lack of knowledge about posttranscriptional contributions to noise by determining cell-to-cell variations in the abundance of mRNA and reporter protein in yeast. Two types of structural element, a stem–loop and a poly(G) motif, not only inhibit translation initiation when inserted into an mRNA 5΄ untranslated region, but also generate noise. The noise-enhancing effect of the stem–loop structure also remains operational when combined with an upstream open reading frame. This has broad significance, since these elements are known to modulate the expression of a diversity of eukaryotic genes. Our findings suggest a mechanism for posttranscriptional noise generation that will contribute to understanding of the generally poor correlation between protein-level stochasticity and transcriptional bursting. We propose that posttranscriptional stochasticity can be linked to cycles of folding/unfolding of a stem–loop structure, or to interconversion between higher-order structural conformations of a G-rich motif, and have created a correspondingly configured computational model that generates fits to the experimental data. Stochastic events occurring during the ribosomal scanning process can therefore feature alongside transcriptional bursting as a source of noise

Engineering Cupriavidus necator H16 for the autotrophic production of (R)-1,3-butanediol

Butanediols are widely used in the synthesis of polymers, specialty chemicals and important chemical intermediates. Optically pure R-form of 1,3-butanediol (1,3-BDO) is required for the synthesis of several industrial compounds and as a key intermediate of β-lactam antibiotic production. The (R)-1,3-BDO can only be produced by application of a biocatalytic process. Cupriavidus necator H16 is an established production host for biosynthesis of biodegradable polymer poly-3-hydroxybutryate (PHB) via acetyl-CoA intermediate. Therefore, the utilisation of acetyl-CoA or its upstream precursors offers a promising strategy for engineering biosynthesis of value-added products such as (R)-1,3-BDO in this bacterium. Notably, C. necator H16 is known for its natural capacity to fix carbon dioxide (CO2) using hydrogen as an electron donor. Here we report engineering of this facultative lithoautotrophic bacterium for heterotrophic and autotrophic production of (R)-1,3-BDO. Implementation of (R)-3-hydroxybutyraldehyde-CoA- and pyruvate-dependent biosynthetic pathways in combination with abolishing PHB biosynthesis and reducing flux through the tricarboxylic acid cycle enabled to engineer strain, which produced 2.97 g/L of (R)-1,3-BDO and achieved production rate of nearly 0.4 Cmol Cmol-1 h-1 autotrophically. This is first report of (R)-1,3-BDO production from CO2

The pMTL70000 modular, plasmid vector series for strain engineering in Cupriavidus necator H16

Cupriavidus necator H16 can convert CO2 into industrial chemicals and fuels. To facilitate its engineering, we designed, built and tested the pMTL70000 modular plasmids comprising standardised Cupriavidus and E. coli replicons, selectable markers and application specific modules. Plasmids were characterised in terms of transmissibility, stability, copy number and compatibility

A genome-wide approach for identification and characterisation of metabolite-inducible systems

© 2020, The Author(s). Inducible gene expression systems are vital tools for the advancement of synthetic biology. Their application as genetically encoded biosensors has the potential to contribute to diagnostics and to revolutionise the field of microbial cell factory development. Currently, the number of compounds of biological interest by far exceeds the number of available biosensors. Here, we address this limitation by developing a generic genome-wide approach to identify transcription factor-based inducible gene expression systems. We construct and validate 15 functional biosensors, provide a characterisation workflow to facilitate forward engineering efforts, exemplify their broad-host-range applicability, and demonstrate their utility in enzyme screening. Previously uncharacterised interactions between sensors and compounds of biological relevance are identified by employing the largest reported library of metabolite-responsive biosensors in an automated high-throughput screen. With the rapidly growing genomic data these innovative capabilities offer a platform to vastly increase the number of biologically detectable molecules