Description

1.0 Introduction

NADH-glutamate dehydrogenase (GDH) is among the most researched enzymes in scientific literature [1], but its physicochemical mechanisms are still evolving [2, 3, 4]. It is the target site of the action of the magnetoelectric changes in the environmental conditions of cells, tissues, and whole organism. GDH captures the changes in the magnetoelectric field by synthesizing nongenetic code-based RNA enzymes (macromolecular NTinti chimerenomic RNA) whose functions are to select and degrade superfluous genetic code-based (mRNA, rRNA, tRNA, long noncoding RNA, chimeric RNA, siRNA, short nuclear RNA etc) RNAs thus preventing them from participating in protein translational processes (please refer to the Table 1 for new terminologies). Chimerenomic (nongenetic code-based) RNAs regulate DNA gene expression; being present in cells and tissues, but in very low quantities [2, 3, 4]. Dysfunctional protein products of superfluous genetic code-based RNAs (mRNA, rRNA, tRNA, circular RNA, long noncoding RNA, chimeric RNA etc) are the root causes of human disease conditions [3].  The thermal stability of chimerenomic RNA compared with total RNA is one of the physicochemical characteristics that enables chimerenomic RNA enzyme to degrade superfluous total RNA [4]; but the structural properties that differentiate the functions of the two kinds of RNA have not been described. Genetic code imposes

Table 1: 1. These new terminologies in life sciences describe the discovery of template-

independent RNA processes that regulate gene expression, and control the survival of cells and organisms, other than genes. The building blocks of this life regulatory system are made up of a unique type of RNA that are not coded nor synthesized through the genetic code. These template-independent RNAs are called chimerenomic RNAs.

2.Chimerenomics: Is the study of everything (NTintis, chigramming, GDH-

chimmerization, chimere etc) about chimerenomic RNAs. Chimerenomics confers molecular chemistry pluripotency and totipotency on all cells and tissues. Chimerenomics are the processes by which whole organisms, cells and tissues differentiate, develop and grow by chigramming chimerenomic RNAs that interact with the changing physicochemical internal and external environmental conditions thereby reprogramming and optimizing those metabolic reactions that assure the continued survival of the organism.

3.Chimere: is the minimum length of nucleotide sequence that can degrade homologous mRNA and other genetic code-based RNAs. Chimere is the active segment of NTinti.

4.NTinti: This is the macromolecular chimerenomic RNA chigrammed (synthesized) by NADH-

glutamate dehydrogenase hexameric redox cycle isoenzymes (GDH) in response to a specific environmental change. NTinti is also synthesized naturally in vivo during normal tissue differentiation, growth and development. Therefore, NTinti can be cell or tissue specific. One NTinti has more than one chimere.

5.Chigramming: This is the process of synthesis of chimeres and NTintis by GDH. It is a

spontaneous process. It is the conversion of the electromagnetic changes in the environmental conditions of cells, tissues, whole organisms to the nucleotide sequences of chimerenomic RNAs.

6.GDH-Chimerization: This is the formation of chemical Schiff base of GDH isoenzymes

in response to a new environment leading to new hexameric isoenzyme complexes. This is the initiation process for chigramming.

7.Functional chimerenomics: is the study of the biological functions of chimere and NTinti.

8.Differential chimerenomics: is the comparison study of the chimeres and NTintis

from same or different tissues under various environmental conditions.

9.Other terminologies

a). Clinical chimerenomics: The application of chimerenomics to clinical studies.

b). Chimerenomic Medicine: This is the application of chimerenomics to primary care in the prevention and treatment of human disorders, diseases and wellness conditions in humans.

c). Chimerenomic Chemistry

d). Chimerenomic Physiology

e). Molecular Chimerenomics

f). Chimerenomic Biology

g). Chimerenomic Biomedical Engineering

h). Chimerenomic Pharmacology

i). Chimerenomic Data Engineering

j). Chimerenomics in Genomics

k). Chimerenomics in Structural Biology etc

steric rigidity [4] on transcripts of DNA gene (mRNA, rRNA, long noncoding RNAs, tRNA etc) to the extent that it affects genetic code-based RNA therapeutics during their formulation as drugs [5]. Chimerenomic RNA’s freedom from steric hindrance has not been described.  Chimerenomics is the new approach in medicine, science, engineering and

biotechnology for conducting drug discovery experiments for the next-generation-drugs. The chimerenomic RNA enzymes chigrammed by human GDHs and the GDHs of the tissues of higher plants have been sequenced.  Chimerenomic RNA enzymes induced by treatment of cultured human astrocyte cells with molar concentrations of narcotic medications are leading the next generation therapies for the potential reversal, treatment and cure of drug addiction including numerous related health disorders [6, 7, 8]. Because chimerenomic medicine and science are stupendous up shoots of contemporary medicine, there is understandably, a vast horizon of new knowledge to ingest in a limited time.  A contemporaneous centralized debacle in the forecourt of the medical disciplines is the need to overhaul the generalized ‘one-disease-one-DNA gene-based’ dogma [5]. As many human disease conditions are in the border line of the inexplicable phenomena in biology, their delayed efficacious treatments are spiraling out of medical control into the zones of ‘disorders of mass destruction’. The multi-dimensional (temporal and spatial) relationships existing between NADH-GDH, chimerenomic RNA enzyme, the multitude of their different homologous genetic code-based RNA targets, and a hodge of protein target sites of disease conditions have opened the flood gate to new multi-modal therapeutics with potential to cure hitherto incurable pandemic-scale human health maladies. The physicochemical mechanism of the degradation of total RNA by chimerenomic RNA is known [2]. But the shape of macromolecular NTinti chimerenomic RNA enzyme has not been described to permit its formulation as drugs by biomedical engineers. With the prevalent promise on chimerenomic science to traverse the fields of medicine, this review strides forward to illuminate a few historical antecedents of the novel RNA and that macromolecular NTinti chimerenomic RNA enzyme is a living molecule with secondary and tertiary structural characteristics favorable for engineering formulation to drugs so that they retain their in vivo capabilities to select and degrade superfluous total RNA.

2.0.Methods
2.1. Treatment of Experimental Organism

Peanut (Arachis hypogea) seeds were sterilized in 5% alcohol solution for 10 min, rinsed with deionized water, and planted on moistened filter paper in five replicate petri dishes. The treatments were:

1. Native 0.1% N-Carboxylmethylchitosan solution (Protein Lab, Redmond, WA, USA);
2. Control: treated with deionized water.

About 8 - 10 seeds were planted per petri dish and allowed to germinate in the greenhouse temperature of 24˚C - 28˚C, and relative humidity of 70% - 80% under Texas month of May temperature and sunlight conditions. The greenhouse was protected 50% with shade cloth. Filter papers were changed and re-wetted with fresh solutions. [9; 10]. After emergence of the full plumule of the control seedlings, germination procedure was stopped; and the seedlings were stored in −30˚C freezer.

2.3. Purification of GDH hexameric isoenzymes

GDH was purified from both the control and the chitosan-treated peanut seedlings (20 g) by homogenization at 4˚C with 100 mL of buffer [11] containing 5 units per mL of each of RNase A, and DNase 1. The homogenate was centrifuged (5000×g, 15 min, 4˚C), and the supernatant was frozen at −80˚C, thawed at room temperature to fracture the mitochondria and to assure that DNA and total RNA have been degraded. After re-centrifugation (9000 ×g, 4˚C, 30 min), the supernatant was subjected to fractional ammonium sulfate precipitation, preparative-scale isoelectric focusing (IEF; Rotofor, Bio-Rad, Hercules, USA) followed by dialysis of the fractions as described before [4]. Rotofor fractions (0.2 mL) were purified by native 7.5 % polyacrylamide gel electrophoresis (PAGE) (100 V, 16 h, 4˚C) to remove other proteins, and nucleic acid contaminations. GDH isoenzymes were eluted (30 min, 100 V) from the electrophoresed gel with 0.05 M solution of Tris base, at subzero temperature in the elution chamber, of mini-whole gel eluter (Bio-Rad) as described before [2]. The eluted GDH charge isomer fractions were collected separately per eluter channel, and stored at −30˚C.

2.4. GDH Chigramming (synthesis) of Chimerenomic (nongenetic code-based) RNA Enzyme.

In the experiments under the pilot projects, all the chimerenomic RNAs were chigrammed in the amination direction in cocktails containing solution of 0.87 mM NH₄Cl, 3.5 mM CaCl₂, 10.0 mM α-ketoglutarate (α-KG), 0.23 mM NADH, 0.6 mM each ribo-NTP, 5 U of RNase inhibitor, 5 U of DNase 1, and 5.0 μg actinomycin D. In the single nucleotide chimerenomic RNAs, each chigramming reaction contained 0.6 mM of a ribo-NTP. In the two-nucleotide chimerenomic RNAs, each chigramming reaction contained 0.6 mM of each of the two ribo-NTPs. In the three-nucleotide chimerenomic RNA reactions, each chigramming reaction contained 0.6 mM of each of the three ribo-NTPs. The control chimerenomic RNA reaction contained 0.6 mM of each of the four ribo-NTPs.  The final volume of each reaction cocktail was brought to 0.4 mL and pH 8 with 0.1 M Tris-HCl solution. The reactions in the single ribo-NTPs, two ribo-NTPs, three ribo-NTPs contained 0.2 mL of cryo-electrophoretically purified chitosan-treated peanut GDH charge isomers about 500 μg protein per mL [12]. Reaction in the control contained 0.2 mL of cryo-electrophoretically purified untreated peanut GDH charge isomers of about 500 µg protein per mL. Reactions were incubated at 16˚C overnight (16 h) and stopped by phenol-chloroform (pH 4.5) extraction of the GDH. Chimerenomic RNA was precipitated with ethanol, and centrifuged at 7000 X g. The slurry liquid chimerenomic RNA was left to dry at room temperature, then suspended in a minimum volume of molecular biology quality water and stored at -70 ^oC in aliquots of 10 µL volumes.  RNA chigramming was repeated three times to verify the reproducibility of the results.

2.5. Primary Structure of Chimerenomic (GDH-synthesized) RNA Enzyme:

Complementary DNAs were synthesized with 2 μg of each product RNA chigrammed by the GDH charge isomers purified from the chitosan-treated peanut seedlings, using random hexamer primers (13). The product cDNA (1 μg) was digested with Taq 1 restriction enzyme (5 Units) for 2 h at 65˚C. Adaptors, ³²P-labeled extension primers, and the selective display PROBE combination (Display Systems Biotech, Vista, CA USA) were ligated to the ends of the restriction fragments. The nucleotide sequences of the adapters, extension primers, and display PROBEs are among the proprietary information of their manufacturer, Display Systems Biotech, Vista, CA USA.

The template DNA (0.5 μg) was amplified according to the ‘touch-down’ restriction fragment double display (RF-DD) PCR methods of Display Systems Biotech, Vista, CA, USA. Initial denaturation was 96˚C for 1 min. For the first 10 cycles: denaturation was at 96˚C for 30 sec., annealing was at 60˚C for 30 sec, for the first cycle, then reduced the annealing temperature 0.5˚C each cycle until an annealing temperature of 55˚C was reached after 10 cycles; extension was at 72˚C for 1 min. PCR was continued another 25 cycles: denaturation (96˚C, 30 sec), annealing (55˚C, 30 sec), extension (72˚C, 1 min); final elongation (72˚C, 5 min). All the 64 display PROBEs in the Display Systems Biotech kit were used in the differential display PCR. The differential bands/products were visualized by autoradiography following polyacrylamide sequencing gel electrophoresis. Selected cDNA fragments were sub-cloned into pCR4-TOPO vector and transformed into TOP10 One Shot Chemically Competent (non-pathogenic) Escherichia coli (Invitrogen, Carlsbad, CA), followed by overnight growth on kanamycin selective plates. Up to 15 positive transformant colonies were picked per plate and cultured overnight in LB medium containing 50 μg/mL of kanamycin.

Plasmids were purified with a plasmid kit (Novagen, Madison, WI), and the insert cDNA fragment of selected recombinant plasmids were sequenced with T7, and T3 primers by MWG Biotech Inc., High Point, North Carolina, USA. For quality control purposes, the sequenced plasmids were amplified by ‘touch-down’ PCR using M13 primers (Invitrogen Life Technologies, California); separated by agarose gel electrophoresis; the product bands were UV-visualized, and photo-documented.

2.6. Secondary and Tertiary Structures of Chimerenomic (GDH-synthesized) RNA Enzyme.

Radio-labeled chimerenomic RNAs were prepared with mixes of 3µL of each of two [α-³²P] NTPs (10 mCi/mL) (ICN). NADH was not added but NH₄Cl (0.8 mM), CaCl₂ (3.0 mM), α-KG (10.0 mM), DNase 1 (5U), RNase inhibitor (5U), and actinomycin D (5 µg) in 0.1 M Tris-HCl buffer (pH 8.0) and about 0.6 µg of GDH were added to bring the final volume to 0.4 mL. Chigramming and precipitation of the chimerenomic liquid RNAs were performed as described above.

            Each radioactive liquid chimerenomic RNA (10 µL) in each microcentrifuge tube was digested with 0.01 Unit each of RNase A, RNase VI, RNase T₁ (Ambion) at room temperature for 15 min according to Ambion’s protocol. There were also controls without RNase. The RNase digestion products were electrophoresed through 1.5% agarose gel, electroblotted (Bio-Rad’s semidry trans-blot cell) onto nylon membrane, followed by autoradiography of the membrane.

3.0 Results and Discussion

3.1. Primary, Secondary, and Tertiary Structures of Chimerenomic RNA Enzymes

All the nucleotide sequences [2, 4, 6, 13, 14] show that the G+C contents of chimerenomic RNA enzymes vary broadly covering a range from 35% to 59%, whereas the G+C contents of mRNAs [4] vary for a narrower range from 50% to 60%. Therefore, chimerenomic RNA enzymes are different from genetic code-based RNA.

The high G+C contents of DNA genes agree with previous observations [4, 15] that the lengths of coding DNA segments in the genomes are under structural constraints associated with their higher G+C contents compared with the lower G+C regions of chimerenomic RNA [4]. But high G+C contents of DNA increase the stability of genes [4]. Previous research on the importance of G+C contents compared the G+C-rich with the G+C-poor regions [4] of genetic code-based DNA. The comparisons between the G+C contents of coding DNA sequence and the chimerenomic RNA enzymes (nongenetic code-based RNA) are at the base line for further unraveling of nucleic acid chemistry and function. Therefore, coding function imposed higher G+C content-related structural constraints on the genome, whereas chimerenomic RNA minimized its G+C content thereby relieving that steric hindrance from its biological function in the degradation of superfluous genetic code-based RNAs and as the universal regulator of gene expression.

The wider range (35% - 59%) in the G+C contents of chimerenomic RNA enzyme compared with the narrower range (50% - 60%) in the genetic code-based RNAs suggests that chimerenomic RNAs possess higher A+T contents which would destabilize at least some local sections in their secondary and tertiary structures. Local helical conformational changes have been reported to exercise biological functions [4]. Such relative differences in the intramolecular A-T and G-C base pairing sections in the NTinti macromolecular chimerenomic RNA enzyme conformations might determine the biological mechanism of the degradation of superfluous genetic code-based RNAs by chimerenomic (nongenetic code-based) RNA enzyme, and the regulation of mRNA abundance through homologous sequence-mediated RNA hydrolytic activity [3, 4, 6, 7]. In describing the features which are correlated with silencing efficiency, Chan et al. [16] identified the importance of low G+C-contents of the siRNA, suggesting that siRNA G+C-content negatively correlated with RNAi efficiency. Liu et al. [17] predicted that efficient siRNAs tend to be A+U-rich (G+C content is 44.0% to 47.8%), whilst inefficient siRNAs tend to be G+C-rich (G+C content is 52.7% to 55.2%).The lower G+C-contents of the  chimerenomic RNAs [4] explain the efficiency of chimerenomic RNA in the degradation of homologous genetic code-based RNAs. The homologous target sequences in the mRNA for the binding of chimerenomic RNA are A+U-rich. Conversely, the higher G+C-contents of genetic code-based RNAs [18] could explain some of their inadequacies in RNA therapeutic technologies [5]. The target sequences in the mRNA for the binding of the genetic code-based siRNA are also G+C-rich. Some publications have stated that the high G+C-contents of genetic code-based siRNA constructs may appear to be the stumbling block in the therapeutic or pharmaceutical application of efficient siRNA technology compared with antisense technology [19, 20]. Chimerenomic RNAs circumvent genetic code-based mRNA structural constraints because GDH implements the chigramming in response to the prevalent metabolic environment of tissues, and cells. We have modified the metabolic environments of several crop species and succeeded in generating chimerenomic RNAs that knocked out specific mRNAs encoding the enzymes for the synthesis of toxic and anti-nutritional components [3, 4, 6, 9]. Also, mammalian GDH chigrams RNA enzymes with high promise to become the next generation of therapeutic drugs for the treatment of environmentally induced human health disorders [3, 6].

The differences between the G+C-contents of DNA gene and chimerenomic RNA [4], and in the amplification uniformity of nongenetic code-based cDNA imply that genetic code-based DNA possesses paramagnetic and electrostatic properties [16] that are different from those of chimerenomic (nongenetic code-based) RNA. Therefore, when chimerenomic RNA enzyme aligns in the correct orientation and interacts with its homologous mRNA (genetic code-based RNA) during degradation reaction, the force of the collusion between the different magntoelectric RNA molecules could be so considerable that the resultant energy could liquefy total RNA, the lesser stable of the two types of RNA [22].

3.2. Ribonuclease Digestion of Chimerenomic RNA Enzymes

NTinti macromolecular chimerenomic RNA enzyme is a single-stranded RNA [3, 6, 13]. The base pairing in NTinti macromolecular chimerenomic RNA enzymes was studied by subjecting the RNAs to digestion with different RNases (ribonucleases).

The chimerenomic RNAs chigrammed by GDH in the presence of two ribo-NTPs had prominent bands in the 26S, 16S, and 5S rRNA molecular mass ranges [13, 14]. All the chimerenomic RNAs containing at least one pyrimidine nucleotide residue were completely degraded by RNase A (Figure 1).

Figure 1: Ribonuclease digestion of chimerenomic RNA enzymes: 1. poly (C,A), 2. poly (U,A), 3. poly (C,G), 4. poly (U,C), and 5. poly (U,G). Each chimerenomic RNA enzyme was digested with (b). RNase A, (c). RNase VI, (d). RNase T₁, (a). Control - no RNase digestion.

RNase VI (hydrolyses at base-paired residues) did not degrade poly (C, A), poly (C, G), poly (U, C), and poly (U, G) chimerenomic RNAs (Figure 1). The lack of base-pairing in the structures of poly (U, C), poly (U, G), and poly (C, A) chimerenomic RNAs is understandable because the U and C, the U and G, and C and A residues did not form stable Watson-Crick base pairing. The lack of base pairs in the poly (C, G) chimerenomic RNA (Figure 1) was surprising but maybe due to a higher frequency of addition of the C than the G by the GDH, thereby producing long stretches of C residues with fewer intermittent G residues in that chimerenomic RNA primary structure.  The observed lower content of G residues in the poly (C, G) chimerenomic RNA was supported by the resistance of that chimerenomic RNA to RNase T₁ (cleaves RNA at purine residues) digestion [13]. The poly (C, G) chimerenomic RNA was almost as resistant to RNase T₁ digestion as those chimerenomic RNAs lacking G residues. Because it lacked any purine residue, the poly (U, C) chimerenomic RNA was the most resistant to RNase T₁ digestion (Figure 1). These results demonstrated the differences in the origins of chimerenomic NTinti macromolecular nongenetic code-based RNA and total RNA (genetic code-based RNA). Chimerenomic RNA is man-made because its primary, secondary, and tertiary structures can be changed at will (Figure 1) depending on the new therapeutic drugs in demand.

3.3. Folded Shape of Macromolecular NTinti Chimerenomic RNA Enzymes:

RNase VI digested the poly (U, A) chimerenomic RNA (Figure 1) completely, probably because the U and A residues were evenly distributed to produce many base-paired regions. The poly (U, A) chimerenomic RNA was also digested to the same extent by RNase A and RNase VI (Figure 1) suggesting that the U residues were as open to attack by RNase A as the U-A base pairs were open to RNase VI digestion. This is proof that the extensive intramolecular base pairing that defines the compact shape (tertiary structures) of chimerenomic RNA enzyme also allows the shape to be folded yet flexible to allow water molecules to be sucked into the intramolecular matrixes so created. These are the structural properties of the liquid chimerenomic RNA enzymes (Figure 1). The presence of extensive intra-molecular base-pairing in the folded structure of macromolecular chimerenomic RNA enzymes portends excellently as a characteristic that could enable the enzyme to form secondary and tertiary compact conformations suitable in the drug’s biomedical engineering formulation procedure. Biological macromolecular chimerenomic RNA enzymes are liquid RNAs, whereas genetic code-based RNAs are solids possessing three-dimensional [23] steric hindrance property. Had their secondary and tertiary structures been as rigid as those of mRNA, long noncoding RNA, rRNA, chimeric RNA, noncoding nuclear RNAs, tRNA etc [24, 25] the drug formulation of the extra-large macromolecular NTinti RNA enzymes would have presented a technological nightmare in biomedical engineering. Active macromolecular NTinti chimerenomic RNA enzymes are the largest man-made biomedical RNA, being as large as 10.5 billion nucleotides long [3, 6, 26], which is almost double the size of human genome.

            Therefore, in the RNase digestion of different chimerenomic RNAs, RNase VI was the main experiment, whilst RNase A was the positive control, and RNase T₁ was the negative control. The poly(U, A) chimerenomic RNA was the main experiment whilst poly(C, A), poly(C, G), poly(U, C), and poly(U, G) chimerenomic RNAs were the controls. So, there were multi-dimensional crisscrossing networks of different RNase and different chimerenomic RNA controls in compliance with the chimerenomic research protocols and practices [2, 3, 6; 7]. The RNase digestions of chimerenomic RNAs revealed unequivocally the base pairing-dependent secondary, and tertiary structures and of the folded shape of chimerenomic RNA enzymes.

3.4. The Pilot Projects

 Several projects were conducted as preface to the selection of the magnetoelectric

environmental conditions for the treatments of seedlings, and for the interpretation of the results of the ribonuclease digestion of the different chimerenomic RNA enzymes to be generated. This was in keeping with the requirement in chimerenomic research protocol for the establishment of a network of negative and positive controls for the delineation of false negatives from the false positives. Chitosan solution was selected as the environmental magnetoelectric field elicitor because it is a biochemical regulator of metabolic processes [27, 28, 29, 30], which property was displayed in the outcomes of the pilot projects (Figure 2). The redox cycle NADH-GDH of the control seedlings without both chitosan solution and ribo-NTP treatment chigrammed the least yield of chimerenomic RNA enzyme in agreement with the role of chitosan in the promotion of growth and differentiation [29, 30].

Figure 2: Pilot project results in selecting the optimal magnetoelectric environmental conditions for the studies on the shape (secondary and tertiary structures) of chimerenomic RNA enzymes. Each single ribo-NTP chimerenomic RNA was chigrammed with 0.6 mM solution of the ribo-NTP; each two ribo-NTPs chimerenomic RNA was chigrammed with 0.6 mM concentration of each ribo-NTP; each three ribo-NTPs chimerenomic RNA was chigrammed with 0.6 mM concentration of each of the three ribo-NTPs, mixed with chigramming reaction cocktail containing NH₄Cl, NADH, α-KG, RNase inhibitor, DNase 1, and actinomycin D and GDH extracted from chitosan-treated peanut seedlings. The control experiment contained 0.6 mM of each of the four ribo-NTPs but the GDH extracted from the control untreated peanut seedlings was used for chigramming. The chimerenomic RNA enzymes produced were ethanol-precipitated, then pelleted by centrifugation, and equal aliquots were applied to agarose gel, and electrophoresed.

            The single ribo-NTP treatments affected the chimerenomic RNA enzyme product yields in the descending order of the nucleophilicities of the nucleotide bases: U > A > C > G (Figure 2), thus explaining the preponderance of A+U contents but low G+C contents of chimerenomic RNAs. This is a major difference between the physicochemical properties of the two kinds of RNAs. It is at this point that genomics diverge from chimerenomics.

The order of the nucleophilicities of the nucleic acid bases explain the differential susceptibilities of the chimerenomic RNA enzymes to the actions of RNase A, RNase VI, and RNase T₁ observed (Figure 1). The mixes of two-NTPs gave similar yields of the chimerenomic RNA, furthermore the mixes of three-NTPs gave similar yields of chimerenomic RNAs; meaning that the different nucleophilicities of ribo-NTPs did not affect the yields of the chimerenomic RNA chigrammed by GDH when different but equal concentration of substrate mixes of ribo-NTPs are provided. However, the primary structures of chimerenomic RNA enzymes differ because the magnetoelectric field under which each was chigrammed was different and unique [2, 3, 6, 7, 9, 10, 13, 14, 21, 22]. The liberation of the genetic code from the primary structure of chimerenomic RNA eliminated the high G+C contents from chimerenomic RNA primary structure. The chimerenomic RNA yields of the two-NTPs mixes were unequivocal in recommending the poly two-NTPs chimerenomics as the pivot of the NTinti chimerenomic RNA shape studies (Figure 1).

The pilot project (Figure 2) embodied many subprojects in different spatial directions, each of which was stratified into multiple layers of experiments to ensure the investigations were quantitative, rigorous, and comprehensive to answer all the intellectual questions to the satisfaction of chimerenomic sciences.    

4.0. Perspectives of Chimerenomic RNA Enzyme Structure and Relevance

Chimerenomic NTinti RNA structure, function, relevance and scientific origin are inseparable.

4.1. The origin of chimerenomic science and medicine research

In 1975, Johnson MW and Osuji GO at the John Innes Center, Norwich UK, made a groundbreaking discovery by developing methods to chemically label, digest, and analyze the base compositions of ribonucleosides and oligonucleotides [31]. Unbeknownst to them, this work was the precursor to a revolutionary branch of nucleic acid research that would evolve into chimerenomic sciences and medicine. Their findings, published in Analytical Biochemistry in 1976 and FEBS Letters in 1977 [31, 32], demonstrated how to determine base composition ratios in sub-microgram quantities of RNA using radioactive p-hydrazinobenzen-[³⁵S] sulphonic acid (³⁵S-p-HBSA). This research expanded the understanding of nucleic acids beyond genomics, highlighting the characteristics of free RNAs derived from viruses, plants, and cells.

Following these discoveries, Osuji continued to investigate these “freelance” RNAs, focusing on their existence, synthesis, roles, and environmental connections separate from genomic studies. Early on, it became evident that stoichiometry dictated the interactions involving nucleoside derivatives, consistently occurring in molar ratios. Thus, these experiments indicated that the engagement of nucleoside derivatives relies on the molar concentrations of ³⁵S-p-HBSA, confirming that chimerenomic reactions happen in specific molar ratios [31, 32].

Soon after the pivotal 1975 experiments, Osuji concluded that two distinct types of RNA exist: one produced through DNA transcription (genetic code-based RNA) and another that is independent of DNA and the genetic code (nongenetic code-based RNA) [8, 31, 32]. RNA viruses are clear examples of naturally occurring RNAs (chimerenomic RNAs) that are not directed by DNA [32]. He later found that higher organisms, including plants, also contain similar RNAs even in the absence of viral infection, leading him to isolate molecules from cells with physicochemical properties like those of these nongenetic code-based RNAs. By the early 1980s, evidence from Osuji’s laboratory suggested that higher organisms could synthesize RNAs independently of the genetic code, and he later showed that this class of RNA (chimerenomic RNA) occurs in small amounts in nearly all higher plant species [3, 13].

From the 1980s to the early 2000s, Osuji research focused on the factors influencing plant growth, examining environmental changes, nutrient availability—including fertilizers and agricultural chemicals as sources of nucleophiles—and their effects on enhancing protein yields as indicators of positive nitrogen balance. This work also highlighted the responsive nature of glutamate dehydrogenase (GDH) isoenzymes to these environmental manipulations, revealing the significant yet previously overlooked role of GDH that had eluded scientists for centuries [21, 27, 28, 29, 30].

This inquiry culminated in the landmark 2004 publication by Osuji GO and colleagues on the RNA synthetic activity of glutamate dehydrogenase [13]. Despite its significance, the scientific community largely overlooked this breakthrough, which demonstrated the capacity to engineer, synthesize and chigram RNAs independent of the genetic code, driven by environmental signals such as nucleophiles and electromagnetic variance.

Chimerenomic sciences emerged in full after the discovery of the RNA synthetic activity of glutamate dehydrogenase (GDH). Starting in 2004, a wealth of insights began to unfold, illuminating this vast and new realm of knowledge. Osuji GO, et al, began to explore the intricacies of chigramming processes and the unique functions and physicochemical properties of this novel type of RNA. This opened infinite possibilities for manipulating environmental factors and their corresponding RNA responses, leading to numerous avenues for exploration and innovation [13, 14].

The discovery that chimerenomic RNAs also function as enzymes [2, 4, 22] marked a significant advancement in the field. It was found that various types of RNA—including mRNA, tRNA, rRNA, siRNA, long non-coding RNA, and chimeric RNA—serve as substrates for chimerenomic RNA. The enzymatic activities of chimerenomic RNAs are not random; rather, they depend on the homology of their substrates to specific segments of the chimerenomic NTinti molecule [22].

Chimerenomic RNAs play a crucial role in protecting cells from environmental disturbances by modulating gene expression, enabling cellular adaptation to occur. In a landmark 2008 publication, Osuji GO and colleagues detailed the RNA synthetic activity of GDH and its implications for drug metabolism research. They demonstrated how nongenetic code-based RNAs derived from GDH isoenzymes could degrade mRNAs that encode various enzymes involved in drug metabolism, including cytochrome P450 reductase, ABC Transporters, and Superoxide dismutase, positioning chimerenomic medicine at the forefront of drug metabolism [14]. This is the emergence and core of chimerenomic medicine.

This work illustrated that chimerenomic RNAs degrade homologous mRNAs derived from the genetic code, a finding consistently validated over the subsequent decade. This led to a deeper understanding of the role of chimerenomic RNA in gene regulation, cellular metabolism and cell function reprogramming [2]. It became evident that environmental changes, often in stoichiometric measures, can trigger cells to activate GDH isoenzymes. If unimpeded, these isoenzymes can chigram chimerenomic RNA, which subsequently regulates gene expression by degrading or modulating genetically encoded RNAs (including mRNA, tRNA, rRNA, siRNA, long noncoding RNA, miRNA and others), thereby influencing protein synthesis and overall cellular metabolism. Ultimately, this reveals that environmental factors primarily dictate gene phenotype expression, reversing the traditional understanding of the gene-product hierarchy.

In 2024, a groundbreaking study by Okea RN and Osuji GO at the American Academy of Primary Care Research, conducted at the Southwest Research Institute in San Antonio, Texas, and sequenced by CD-Genomics in New York, solidified the foundations of chimerenomic medicine. This research challenged existing dogmas in medical and drug research, introducing a novel methodology for drug discovery and treatment of diseases, thus establishing chimerenomic medicine as the future of medicine [3, 6, 7, 26].

Through chimerenomic techniques, the AAPCR team pioneered research on human astrocyte cells, demonstrating that clinical research and drug development could directly emerge from chimerenomic-cell-pharmacoactive-macromolecular-NTinti-enzyme systems.

The meticulously engineered and chigrammed molecules exhibited direct applicability to human studies, establishing an expedited pathway for drug discovery and disease treatment.

The study revealed that a pharmaco-active signal could provoke chimerenomic RNA chigramming in accordance with its molar concentration. As the molar concentration varied, different chimerenomic RNAs were generated [3, 6, 7, 26] This stoichiometric response in chimerenomic RNA chigramming is a unique characteristic of chimerenomic experimentation, highlighting that specific molar concentrations can yield optimal results in NTinti RNA enzyme production.

This study culminated in the creation of the largest synthetic macromolecule to date: a chimerenomic NTinti RNA enzyme comprising 10.5 billion nucleotides, generated after human astrocyte cells were exposed to various active pharmaceutical agents under controlled environmental conditions. These innovative molecules demonstrated in vitro capabilities in homology testing, effectively degrading homologous mRNAs linked to several human diseases, including Alzheimer’s and Parkinson’s disease [3, 6].

A comprehensive database of chimerenomic RNA is now accessible to researchers at www.chimerenomics.com. Additionally, the introduction of the chimerenomic pump allows for the synthesis and chigramming of nongenetic code-based chimerenomic macromolecular NTinti RNAs, which can contain billions of nucleotide bases with infinite combinations. This ability to manipulate environmental interventions means that we can accurately determine the nucleoside base compositions of NTinti RNA and specifically tailor it to cure specific diseases [2, 3, 6, 7, 26].

The vast potential of chimerenomic science and medicine represents an ever-expanding landscape of possibilities, perhaps even surpassing the capabilities offered by AI technology.

One of the notable achievements in chimerenomic sciences and medicine is the creation of the chimerenomic search engine [3, 6], a comprehensive database of chimerenomic NTinti macromolecules funded by the American Academy of Primary Care Research (AAPCR). This database surpasses the size of the NCBI Nucleotide BLAST and offers limitless possibilities and potential. Unlike the static BLAST system, the chimerenomic database is dynamic, continuously adapting to reflect changing environmental factors, modulations, and compares of cellular responses across various conditions.

The chimerenomic search engine also serves as a valuable tool for direct clinical studies and drug discovery, featuring a built-in homology comparison tool [3, 6, 7, 26]. This enables research scientists to assess the homology of NTinti RNAs against target mRNAs before initiating NTinti RNA drug production. As a result, NTinti drug derivatives can be identified even prior to clinical trials, with these trials serving simply to validate the already established target effects. By combining the capabilities of the chimerenomic search tool with those of the BLAST system, research scientists and pharmaceutical industries are presented with unprecedented opportunities for drug discovery and development [2, 3, 6, 7, 26].

4.2. Properties of Macromolecular NTinti RNA Drugs

Chimerenomic medicine has provided insights into the various properties of macromolecular NTinti RNA drugs, addressing critical aspects such as structure, solubility, formulation, selectivity, immunogenicity, and delivery to target organs.

As discussed in Section 3, the primary, secondary, and tertiary structures of nongenetic code-based macromolecular NTinti RNAs are firmly grounded in nucleic acid chemistry. This research highlights a distinct contrast between genetic code-based RNAs, like mRNA, and non-genetic code-based RNAs, such as chimerenomic NTinti RNA.

Key differences include:

• G+C Ratio: Genetic code-based RNAs typically exhibit a higher and narrower G+C ratio (50% to 60%), while NTinti RNAs show a lower and broader G+C ratio (35% to 59%) [4].
• Steric Hindrance: mRNAs are affected by genetic code-imposed steric hindrance, limiting their structural flexibility. In contrast, NTinti RNAs do not experience steric hindrance, allowing for greater adaptability.
• Solubility: mRNAs are solid and sparingly soluble, whereas NTinti RNAs are liquid and exceptionally soluble. This solubility makes NTinti RNAs ideal candidates for pharmacological formulations.

For instance, a mere 1 microliter (1 µL) of NTinti RNA can contain over 10.5 billion nucleotides, facilitating easy formulation into liquid nanoparticles (LNP) for efficient drug delivery [2, 3, 6, 7, 26]. These unique properties enhance the potential of NTinti RNAs in therapeutic applications, making them preferable candidates for developing advanced drug delivery systems. NTinti RNAs are species-specific, meaning that those intended for human use must be synthesized from human cells, while NTinti RNAs intended for rats should be produced from rat cells, and so forth. They are also tissue-specific in their actions; NTinti RNA from one tissue will degrade total RNA from that same tissue [4, 8, 22]. Importantly, nucleotides themselves are not immunogenic and by extension NTinti RNAs; they do not provoke an immune response or the production of antibodies on their own [33, 34]. This characteristic further supports the safety and compatibility of NTinti RNA drugs within the same species, enhancing their therapeutic potential. In practice, chimerenomic NTinti RNAs are generated by mimicking the natural adaptive mechanisms of the target host cells. As a result, the host is more likely to recognize them as “self” RNA, which reduces the likelihood of immunogenicity [34, 35, 36, 37].

Table 2.

The differences between chimerenomic NTinti RNA and genetic code-based RNA

NTinti RNAs can degrade abnormal homologous total RNAs and mRNAs [2, 22]. This essential property allows for selective targeting in therapeutics, enabling treatment that minimizes unexpected side effects and avoids unintended clinical complications. By specifically targeting only the aberrant RNA sequences, NTinti RNAs enhance the precision of therapeutic interventions, making them valuable tools in modern medicine.