Beyond a Quartic Polynomial Modeling of the DNA Double-Helix Genetic Code

By combination of finite number theory and quantum information, the complete quantum information in the DNA genetic code has been made likely by Planat et al. (2020). In the present contribution a varied quartic polynomial contrasting the polynomial used by Planat et al. is proposed that considered apart from the golden mean also the fifth power of this dominant number of nature to adapt the code information. The suggested polynomial is denoted as g(x) = x4 - x3 - (4 - ϕ2 )x2 + (4 – ϕ2)x + 1, where is the golden mean. Its roots are changed to more golden mean based ones in comparison to the Planat polynomial. The new coefficients 4 – ϕ2 instead of 4 would implement the fifth power of the golden mean indirectly applying . As an outlook, it should be emphesized that the connection between genetic code and resonance code of the DNA may lead us to a full understanding of how nature stores and processes compacted information and what indeed is consciousness linking everything with each other suggestedly mediated by all-pervasive dark constituents of matter respectively energy. The number-theoretical approach to DNA coding leads to the question about the helical structure of the electron.

Applications of genetic code expansion technology in eukaryotes

Unnatural amino acids(UAAs)have gained significant attention in protein engineering and drug development owing to their ability to introduce new chemical functionalities to proteins.In eukaryotes,genetic code expansion(GCE)enables the incorporation of UAAs and facilitates posttranscriptional modification(PTM),which is not feasible in prokaryotic systems.GCE is also a powerful tool for cell or animal imaging,the monitoring of protein interactions in target cells,drug development,and switch regulation.Therefore,there is keen interest in utilizing GCE in eukaryotic systems.This review provides an overview of the application of GCE in eukaryotic systems and discusses current challenges that need to be addressed.

A Content-Centric Organization of the Genetic Code

The codon table for the canonical genetic code can be rearranged in such a way that the code is divided into four quarters and two halves according to the variability of their GC and purine contents, respectively. For prokaryotic genomes, when the genomic GC content increases, their amino acid contents tend to be restricted to the GC-rich quarter and the purine-content insensitive half, where all codons are fourfold degenerate and relatively mutation-tolerant. Conversely, when the genomic GC content decreases, most of the codons retract to the AUrich quarter and the purine-content sensitive half; most of the codons not only remain encoding physicochemically diversified amino acids but also vary when transversion （between purine and pyrimidine） happens. Amino acids with sixfolddegenerate codons are distributed into all four quarters and across the two halves; their fourfold-degenerate codons are all partitioned into the purine-insensitive half in favorite of robustness against mutations. The features manifested in the rearranged codon table explain most of the intrinsic relationship between protein coding sequences （the informational content） and amino acid compositions （the functional content）. The renovated codon table is useful in predicting abundant amino acids and positioning the amino acids with related or distinct physicochemical properties.

A Scenario on the Stepwise Evolution of the Genetic Code

It is believed that in the RNA world the operational （ribozymes） and the informational （riboscripts） RNA molecules were created with only three （adenosine, uridine, and guanosine） and two （adenosine and uridine） nucleosides, respectively, so that the genetic code started uncomplicated. Ribozymes subsequently evolved to be able to cut and paste themselves and riboscripts were acceptive to rigorous editing （adenosine to inosine）; the intensive diversification of RNA molecules shaped novel cellular machineries that are capable of polymerizing amino acids-a new type of cellular building materials for life. Initially, the genetic code, encoding seven amino acids, was created only to distinguish purine and pyrimidine; it was later expanded in a stepwise way to encode 12, 15, and 20 amino acids through the relief of guanine from its roles as operational signals and through the recruitment of cytosine. Therefore, the maturation of the genetic code also coincided with （1） the departure of aminoacyl-tRNA synthetases （AARSs） from the primordial translation machinery, （2） the replacement of informational RNA by DNA, and （3） the co-evolution of AARSs and their cognate tRNAs. This model predicts gradual replacements of RNA-made molecular mechanisms, cellular processes by proteins, and informational exploitation by DNA.

Therapeutic applications of genetic code expansion

In nature,a limited,conservative set of amino acids are utilized to synthesize proteins.Genetic code expansion technique reassigns codons and incorporates noncanonical amino acids(ncAAs)through orthogonal aminoacyltRNA synthetase(aaRS)/tRNA pairs.The past decade has witnessed the rapid growth in diversity and scope for therapeutic applications of this technology.Here,we provided an update on the recent progress using genetic code expansion in the following areas:antibody-drug conjugates(ADCs),bispecific antibodies(BsAb),immunotherapies,long-lasting protein therapeutics,biosynthesized peptides,engineered viruses and cells,as well as other therapeutic related applications,where the technique was used to elucidate the mechanisms of biotherapeutics and drug targets.

On the Organizational Dynamics of the Genetic Code

The organization of the canonical genetic code needs to be thoroughly illuminated. Here we reorder the four nu- cleotides--adenine, thymine, guanine and cytosine--according to their emergence in evolution, and apply the or- ganizational rules to devising an algebraic representation for the canonical genetic code. Under a framework of the devised code, we quantify codon and amino acid usages from a large collection of 917 prokaryotic genome sequences, and associate the usages with its intrinsic structure and classification schemes as well as amino acid physicochemical properties. Our results show that the algebraic representation of the code is structurally equiva- lent to a content-centric organization of the code and that codon and amino acid usages under different classifica- tion schemes were correlated closely with GC content, implying a set of rules governing composition dynamics across a wide variety of prokaryotic genome sequences. These results also indicate that codons and amino acids are not randomly allocated in the code, where the six-fold degenerate codons and their amino acids have important balancing roles for error minimization. Therefore, the content-centric code is of great usefulness in deciphering its hitherto unknown regularities as well as the dynamics of nucleotide, codon, and amino acid compositions.

Does the Genetic Code Have A Eukaryotic Origin?

In the RNA world, RNA is assumed to be the dominant macromolecule performing most, if not all, core ＂house-keeping＂ functions. The ribo-cell hypothesis suggests that the genetic code and the translation machinery may both be born of the RNA world, and the introduction of DNA to ribo-cells may take over the informational role of RNA gradually, such as a mature set of genetic code and mech- anism enabling stable inheritance of sequence and its variation. In this context, we modeled the genetic code in two content variables^C and purine contents--of protein-coding sequences and measured the purine content sensitivities for each codon when the sensitivity （% usage） is plotted as a function of CJC content variation. The analysis leads to a new pattern--the symmetric pattern--where the sensitivity ofpurine content variation shows diagonally symmetry in the codon table more significantly in the two GC content invariable quarters in addition to the two existing patterns where the table is divided into either four GC content sensitivity quarters or two amino acid diversity halves. The most insensitive codon sets are GUN （valine） and CAN （CAR for asparagine and CAY for aspartic acid） and the most biased amino acid is valine （always over-estimated） followed by alanine （always under-estimated）. The unique position of valine and its codons suggests its key roles in the final recruitment of the complete codon set of the canonical table. The distinct choice may only be attributable to sequence signatures or signals of splice sites for spliceosomal introns shared by all extant eukaryotes.

Prebiotic Chemistry in Aqueous Environment:A Review of Peptide Synthesis and Its Relationship with Genetic Code

Prebiotic peptide synthesis and the origin of the genetic code are central issues concerning the origin of life.The question of how they are possibly correlated on the primordial Earth remains perplexing,although numerous experiments have been carried out to explain the prebiotic chemistry of peptide synthesis and the genetic code origin.The purpose of this article is to review the chemical reactions occurred during the synthesis of peptides and the origin of the genetic code in the early Earth aqueous environment.Meanwhile,we attempt to review their relationship as well.At last,from our perspective,the chiral properties of biomolecules should be taken into account in the prebiotic chemical scenarios,which may contribute to some breakthroughs in the further research of this field.

Dynamic airspace sectorization via improved genetic algorithm

This paper deals with dynamic airspace sectorization （DAS） problem by an improved genetic algorithm （iGA）. A graph model is first constructed that represents the airspace static structure. Then the DAS problem is formulated as a graph-partitioning problem to balance the sector workload under the premise of ensuring safety. In the iGA, multiple populations and hybrid coding are applied to determine the optimal sector number and airspace sectorization. The sector constraints are well satisfied by the improved genetic operators and protect zones. This method is validated by being applied to the airspace of North China in terms of three indexes, which are sector balancing index, coordination workload index and sector average flight time index. The improvement is obvious, as the sector balancing index is reduced by 16.5 %, the coordination workload index is reduced by 11.2 %, and the sector average flight time index is increased by 11.4 % during the peak-hour traffic.

Light-controlled phosphorylation in the TrkA-Y785 site by photosensitive UAAs activates the MAPK/ERK signaling pathway

Background:This paper aims to establish a light-controlled phosphorylation detection method at the Y785 site of tropomyosin receptor kinase A(TrkA)receptor in mammalian cells by using genetic code expansion technology and detecting the effects of optical activation of this site on the downstream MAPK/ERK pathway.The study is based on the current situation that the regulatory mechanism of TrkA phosphorylation has not been fully elucidated.Methods:Two photosensitive unnatural amino acids,p-azido-L-phenylalanine(AzF)and photo-caged tyrosine(ONB)were introduced into the TrkA-Y785 site by genetic code expansion technology and site-directed mutagenesis.Western blotting and laser confocal imaging were conducted to analyze the effects of this site on activating the MAPK/ERK pathway and nerve cell differentiation before and after photostimulation.Results:Our results supplemented the light-controlled results of the TrkA-Y785 site based on our previous research and verified that Y785 also makes important contributions in regulating the MAPK/ERK pathway.Conclusion:This study demonstrated the significant contributions of the TrkAY785 site in regulating the ERK pathway by precisely controlling the phosphorylation state of a single tyrosine site.

Optimization of Process Parameters of Continuous Microwave Drying Raspberry Puree Based on RSM and ANN-GA

To improve drying uniformity and anthocyanin content of the raspberry puree dried in a continuous microwave dryer,the effects of process parameters(microwave intensity,air velocity,and drying time)on evaluation indexes(average temperature,average moisture content,average retention rate of the total anthocyanin content,temperature contrast value,and moisture dispersion value)were investigated via the response surface method(RSM)and the artificial neural network(ANN)with genetic algorithm(GA).The results showed that the microwave intensity and drying time dominated the changes of evaluation indexes.Overall,the ANN model was superior to the RSM model with better estimation ability,and higher drying uniformity and anthocyanin retention rate were achieved for the ANN-GA model compared with RSM.The optimal parameters were microwave intensity of 5.53 W•g^(-1),air velocity of 1.22 m·s^(-1),and drying time of 5.85 min.This study might provide guidance for process optimization of microwave drying berry fruits.

Site-specific protein modification by genetic encoded disulfide compatible thiols

Cysteine chemistry provides a low cost and convenient way for site-specific protein modification.However,recombinant expression of disulfide bonding containing protein with unpaired cysteine is technically challenging and the resulting protein often suffers from significantly reduced yield and activity.Here we used genetic code expansion technique to introduce a surface exposed self-paired dithiol functional group into proteins,which can be selectively reduced to afford active thiols.Two compounds containing self-paired disulfides were synthesized,and their genetic incorporations were validated using green fluorescent proteins(GFP).The compatibility of these self-paired di-thiols with natural disulfide bond was demonstrated using antibody fragment to afford site-specifically labeled antibody.This work provides another valuable building block into the chemical tool-box for site-specific labeling of proteins containing internal disulfides.

Progress of Research on Origins of Life in China

The development of Chinese space science and technology plays a great role in promoting the researches in the field of the origin of life.With the multidisciplinary cooperation,there are fruitful achievements in this research field obtained over the past two years.This report summarizes the major progress of the basic researches about the origin of life in China during 2018–2020.

Genetic Encoding of a Photocaged Glutamate for Optical Control of Protein Functions

Genetic encoding of photocaged noncanonical amino acids provides a powerful tool to study protein functions through optical control but is not yet available for acidic amino acids.Herein,we report the first site-specific genetic encoding of a photocaged glutamate,4-methoxy-7-nitroindolinyl caged glutamate(MNI-Glu),into recombinant proteins via an expanded genetic code through evolved EcLeuRS/tRNA pair.Using two enzymes as examples,we demonstrate that substituting the conserved-active-site glutamate of a secreted alkaline phosphatase and a protease HRV3C to MNI-Glu allows photoregulatory control of their enzymatic activities.Our approach is an important addition to the photocaged noncanonical amino-acid toolbox and provides a general method to photocontrol protein activity based on caging a critical glutamate.

Site-specific incorporation of reduction-controlled guest amino acids into proteins for cucurbituril recognition

Protein recognition using host-guest recognition approach is of great interest but has been limited mainly to the protein N-terminal residues.Here,we site-specific incorporated two novel non-canonical amino acids containing supramolecular guest motifs into protein via an expanded genetic code.Through Staudinger reduction reactions,the encoded unnatural residues on protein becoming activated and can be specifically recognized by cucurbit[7]uril(CB[7])and cucurbit[8]uril(CB[8]).We demonstrated that enzyme containing guest amino acid incorporated near the active site can be reversibly regulated by CB[7]recognition,and CB[8]recognition induces protein dimerization.These amino acids will make useful addition to the supramolecular toolbox for protein targeting using molecular recognition approaches.

A Method of Emergency Volunteer Team Internal Participation in Rescue Decision-Making Considering Psychological Behavior

A method for internal participation in rescue decision-making of emergency volunteer teams considering psychological behavior is proposed to address the time sequence of rescue tasks.Firstly,the problem of multi-tasking and multi-operation within the emergency volunteer team is described.Secondly,considering that task leaders are influenced by behavioral and psychological factors in the evaluation,the required time for the job is used as a reference point,and the expected time that volunteers can complete the job is used as an attribute value.The task leader's prospect satisfaction value for each volunteer is calculated based on prospect theory,and the perceived utility values of disappointment theory and regret theory are calculated to measure the task leader's satisfaction with each volunteer.Furthermore,a multilayer coded genetic algorithm is used to construct an optimization model for emergency volunteer decision-making with the objective of maximizing the satisfaction value.Finally,the feasibility and effectiveness of this method are illustrated by an example analysis.The result shows that the efficiency of rescue tasks can be improved through decision optimization within the volunteer team.

Thirty Years of Multiple Sequence Codes

An overview is presented on the status of studies on multiple codes in genetic sequences. Indirectly, the existence of multiple codes is recognized in the form of several rediscoveries of Second Genetic Code that is different each time. A due credit is given to earlier seminal work related to the codes often neglected in literature. The latest developments in the field of chromatin code are discussed, as well as perspectives of single-base resolution studies of nucleosome positioning, including rotational setting of DNA on the surface of the histone octamers.

Simplified methodology for a modular and genetically expanded protein synthesis in cell-free systems

Genetic code expansion,which enables the site-specific incorporation of unnatural amino acids into proteins,has emerged as a new and powerful tool for protein engineering.Currently,it is mainly utilized inside living cells for a myriad of applications.However,the utilization of this technology in a cell-free,reconstituted platform has several advantages over living systems.The typical limitations to the employment of these systems are the laborious and complex nature of its preparation and utilization.Herein,we describe a simplified method for the preparation of this system from Escherichia coli cells,which is specifically adapted for the expression of the components needed for cell-free genetic code expansion.Besides,we propose and demonstrate a modular approach to its utilization.By this approach,it is possible to prepare and store different extracts,harboring various translational components,and mix and match them as needed for more than four years retaining its high efficiency.We demonstrate this with the simultaneous incorporation of two different unnatural amino acids into a reporter protein.Finally,we demonstrate the advantage of cell-free systems over living cells for the incorporation ofδ-thio-boc-lysine into ubiquitin by using the methanosarcina mazei wild-type pyrrolysyl tRNACUA and tRNA-synthetase pair,which could not be achieved in a living cell.

The Pendulum Model for Genome Compositional Dynamics:from the Four Nucleotides to the Twenty Amino Acids

The genetic code serves as one of the natural links for life＇s two conceptual frameworks--the informational and operational tracks-- bridging the nucleotide sequence of DNA and RNA to the amino acid sequence of protein and thus its structure and function. On the informational track, DNA and its four building blocks have four basic variables： order, length, GC and purine contents; the latter two exhibit unique characteristics in prokaryotic genomes where protein-coding sequences dominate. Bridging the two tracks, tRNAs and their aminoacyl tRNA synthases that interpret each codon--nucleotide triplet, together with ribosomes, form a complex machinery that translates genetic information encoded on the messenger RNAs into proteins. On the operational track, proteins are selected in a context of cellular and organismal functions constantly. The principle of such a functional selection is to minimize the damage caused by sequence alteration in a seemingly random fashion at the nucleotide level and its function-altering consequence at the protein level; the principle also suggests that there must be complex yet sophisticated mechanisms to protect molecular interactions and cellular processes for cells and organisms from the damage in addition to both immediate or short-term eliminations and long-term selections. The two- century study of selection at species and population levels has been leading a way to understand rules of inheritance and evolution at molecular levels along the informational track, while ribogenomics, epigenomics and other operationally-defined omits （such as the metabolite-centric metabolomics） have been ushering biologists into the new millennium along the operational track.

Local Conformational Constraint of Firefly Luciferase Can Affect the Energy of Bioluminescence and Enzyme Stability

Conformational dynamics contribute importantly to enzyme catalysis,such that targeted conformational constraint may affect catalysis.Firefly luciferases undergo extensive structural change during catalysis;key residues form a hydrophobic pocket,excluding water and enabling maximally energetic light production.Point mutants almost always luminesce at longer wavelengths(lower energy)than the wild type.Conformational constraint,using dipeptide analogue 3 at a position critical for optimized excited state structure,produced luciferase emission at a shorter wavelength by∼10 nm.Incomparison,introduction of conformationally constrained analogues 4,5,or 7 afforded luciferases emitting at longer wavelengths,while a related unconstrained luciferase(analogue 6)exhibited wild-type emission.The constrained luciferases tested were more stable than the wild type.Protein modeling demonstrated that the“inside”or“outside”orientation of the conformationally constrained dipeptide led to the shorter or longer emission wavelength,respectively.More broadly,these results suggest that local conformational constraint can control specific elements of enzyme behavior,both in vitro and in vivo.This represents the first example of studying enzyme function by introducing conformationally constrained dipeptides at a specific protein position.The principles discovered here in luciferase modification will enable studies to control the wavelength emission and photophysical properties of modified luciferases.