Making Sense of “Nonsense” and More: Challenges and Opportunities in the Genetic Code Expansion, in the World of tRNA Modifications

The evolutional development of the RNA translation process that leads to protein synthesis based on naturally occurring amino acids has its continuation via synthetic biology, the so-called rational bioengineering. Genetic code expansion (GCE) explores beyond the natural translational processes to further enhance the structural properties and augment the functionality of a wide range of proteins. Prokaryotic and eukaryotic ribosomal machinery have been proven to accept engineered tRNAs from orthogonal organisms to efficiently incorporate noncanonical amino acids (ncAAs) with rationally designed side chains. These side chains can be reactive or functional groups, which can be extensively utilized in biochemical, biophysical, and cellular studies. Genetic code extension offers the contingency of introducing more than one ncAA into protein through frameshift suppression, multi-site-specific incorporation of ncAAs, thereby increasing the vast number of possible applications. However, different mediating factors reduce the yield and efficiency of ncAA incorporation into synthetic proteins. In this review, we comment on the recent advancements in genetic code expansion to signify the relevance of systems biology in improving ncAA incorporation efficiency. We discuss the emerging impact of tRNA modifications and metabolism in protein design. We also provide examples of the latest successful accomplishments in synthetic protein therapeutics and show how codon expansion has been employed in various scientific and biotechnological applications.

Fault detection and diagnosis in building energy systems: A tool chain for the automated generation of training data

The application of fault detection and diagnosis (FDD) algorithms in building energy management systems (BEMS) has great potential to increase the efficiency of building energy systems (BES). The usage of supervised learning algorithms requires time series depicting both nominal and component faulty behaviour for their training. In this paper, we introduce a method that automates Modelica code extension of BES models in Python with fault models to approximate real component faults. The application shows two orders of magnitude faster implementation compared to manual modelling, while no errors occur in the connections between fault and component models.

Some theoretical aspects of reprogramming the standard genetic code

1AbstractReprogramming of the standard genetic code in order to include non-canonical amino acids (ncAAs) opens a new perspective in medicine, industry and biotechnology. There are several methods of engineering the code, which allow us for storing new genetic information in DNA sequences and transmitting it into the protein world. Here, we investigate the problem of optimal genetic code extension from theoretical perspective. We assume that the new coding system should encode both canonical and new ncAAs using 64 classical codons. What is more, the extended genetic code should be robust to point nucleotide mutation and minimize the possibility of reversion from new to old information. In order to do so, we follow graph theory to study the properties of optimal codon sets, which can encode 20 canonical amino acids and stop coding signal. Finally, we describe the set of vacant codons that could be assigned to new amino acids. Moreover, we discuss the optimal number of the newly incorporated ncAAs and also the optimal size of codon blocks that are assigned to ncAAs.

Basic principles of the genetic code extension

Compounds including non-canonical amino acids (ncAAs) or other artificially designed molecules can find a lot of applications in medicine, industry and biotechnology. They can be produced thanks to the modification or extension of the standard genetic code (SGC). Such peptides or proteins including the ncAAs can be constantly delivered in a stable way by organisms with the customized genetic code. Among several methods of engineering the code, using non-canonical base pairs is especially promising, because it enables generating many new codons, which can be used to encode any new amino acid. Since even one pair of new bases can extend the SGC up to 216 codons generated by a six-letter nucleotide alphabet, the extension of the SGC can be achieved in many ways. Here, we proposed a stepwise procedure of the SGC extension with one pair of non-canonical bases to minimize the consequences of point mutations. We reported relationships between codons in the framework of graph theory. All 216 codons were represented as nodes of the graph, whereas its edges were induced by all possible single nucleotide mutations occurring between codons. Therefore, every set of canonical and newly added codons induces a specific subgraph. We characterized the properties of the induced subgraphs generated by selected sets of codons. Thanks to that, we were able to describe a procedure for incremental addition of the set of meaningful codons up to the full coding system consisting of three pairs of bases. The procedure of gradual extension of the SGC makes the whole system robust to changing genetic information due to mutations and is compatible with the views assuming that codons and amino acids were added successively to the primordial SGC, which evolved minimizing harmful consequences of mutations or mistranslations of encoded proteins.

Basic principles of the genetic code extension

Compounds including non-canonical amino acids or other artificially designed molecules can find a lot of applications in medicine, industry and biotechnology. They can be produced thanks to the modification or extension of the standard genetic code (SGC). Such peptides or proteins including the non-canonical amino acids can be constantly delivered in a stable way by organisms with the customized genetic code. Among several methods of engineering the code, using non-canonical base pairs is especially promising, because it enables generating many new codons, which can be used to encode any new amino acid. Since even one pair of new bases can extend the SGC up to 216 codons generated by six-letter nucleotide alphabet, the extension of the SGC can be achieved in many ways. Here, we proposed a stepwise procedure of the SGC extension with one pair of non-canonical bases to minimize the consequences of point mutations. We reported relationships between codons in the framework of graph theory. All 216 codons were represented as nodes of the graph, whereas its edges were induced by all possible single nucleotide mutations occurring between codons. Therefore, every set of canonical and newly added codons induces a specific subgraph. We characterized the properties of the induced subgraphs generated by selected sets of codons. Thanks to that, we were able to describe a procedure for incremental addition of the set of meaningful codons up to the full coding system consisting of three pairs of bases. The procedure of gradual extension of the SGC makes the whole system robust to changing genetic information due to mutations and is compatible with the views assuming that codons and amino acids were added successively to the primordial SGC, which evolved to minimize harmful consequences of mutations or mistranslations of encoded proteins.

pyKNEEr: An image analysis workflow for open and reproducible research on femoral knee cartilage

Transparent research in musculoskeletal imaging is fundamental to reliably investigate diseases such as knee osteoarthritis (OA), a chronic disease impairing femoral knee cartilage. To study cartilage degeneration, researchers have developed algorithms to segment femoral knee cartilage from magnetic resonance (MR) images and to measure cartilage morphology and relaxometry. The majority of these algorithms are not publicly available or require advanced programming skills to be compiled and run. However, to accelerate discoveries and findings, it is crucial to have open and reproducible workflows. We presentpyKNEEr, a framework for open and reproducible research on femoral knee cartilage from MR images.pyKNEEris written in python, uses Jupyter notebook as a user interface, and is available on GitHub with a GNU GPLv3 license. It is composed of three modules: 1) image preprocessing to standardize spatial and intensity characteristics, 2) femoral knee cartilage segmentation for intersubject, multimodal, and longitudinal acquisitions, and 3) analysis of cartilage morphology and relaxometry. Each module contains one or more Jupyter notebooks with narrative, code, visualizations, and dependencies to reproduce computational environments.pyKNEErfacilitates transparent image-based research of femoral knee cartilage because of its ease of installation and use, and its versatility for publication and sharing among researchers. Finally, due to its modular structure,pyKNEErfavors code extension and algorithm comparison. We tested our reproducible workflows with experiments that also constitute an example of transparent research withpyKNEEr. We provide links to executed notebooks and executable environments for immediate reproducibility of our findings.

Considerations of Alloy N Code Extension for Commercial Molten Salt Reactor Development and Deployment

For commercial development and deployment of the molten salt reactor, a structural alloy that provides both strength at high temperature and resistance to very corrosive molten salt environment is required. To meet this requirement, a survey is conducted on domestic and international candidate alloys. Alloy N turns out to be the sole frontrunner in readiness for qualification to enable the desired deployment within an estimated 10 years. A review of the qualification for commercial nuclear applications indicates that Alloy N has met a large portion of the requirements. Gaps in the qualification are also identified. A search for historical data is underway to retrieve information needed for filling the gaps and upgrading the qualification. Scope of the discovered historical data is briefly discussed and strategic planning for research and development pathway is suggested to ensure successful evolution in commercial deployment of the molten salt reactor system.