Genomic Intelligence as Über Bio-Cybersecurity: The Gödel Sentence in Immuno-Cognitive Systems

This paper gives formal foundations and evidence from gene science in the post Barbara McClintock era that the Gödel Sentence, far from being an esoteric construction in mathematical logic, is ubiquitous in genomic intelligence that evolved with multi-cellular life. Conditions uniquely found in the Adaptive Immune System (AIS) and Mirror Neuron System (MNS), termed the genomic immuno-cognitive system, coincide with three building blocks in computation theory of Gödel, Turing and Post (G-T-P). (i) Biotic elements have unique digital identifiers with gene codes executing 3D self-assembly for morphology and regulation of the organism using the recursive operation of Self-Ref (Self-Reference) with the other being a self-referential projection of self. (ii) A parallel offline simulation meta/mirror environment in 1–1 relation to online machine executions of self-codes gives G-T-P Self-Rep (Self-Representation). (iii) This permits a digital biotic entity to self-report that it is under attack by a biotic malware or non-self antigen in the format of the Gödel sentence, resulting in the “smarts” for contextual novelty production. The proposed unitary G-T-P recursive machinery in AIS and in MNS for social cognition yields a new explanation that the Interferon Gamma factor, known for friend-foe identification in AIS, is also integral to social behaviors. New G-T-P bio-informatics of AIS and novel anti-body production is given with interesting testable implications for COVID-19 pathology.     

Ultrasensitive detection of indoleamines by combination of nanoparticle-based extraction with capillary electrophoresis/laser-induced native fluorescence

For the first time, citrate-capped gold nanoparticles (citrate-AuNPs) have been used for the selective extraction of indoleamines – 5-hydroxytryptophan (5-HTP), tryptophan (Trp), tryptamine (TA), 5-hydroxytryptamine (5-HT), and 5-hydroxyindoleacetic acid (5-HIAA) – prior to their analysis by capillary electrophoresis/laser-induced native fluorescence (CE/LINF). The extinction spectra obtained for the citrate-AuNPs in the presence of indoleamines revealed that 5-HTP, 5-HT, and 5-HIAA were extracted mainly because of van der Waals interactions between the indole ring and the citrate-AuNPs (hydrophobic surface), while 5-HT and TA were extracted by electrostatic attractions between the amine group of the indoleamines and the citrate ligands adsorbed on the AuNP surface. The extracted indoleamines could be liberated from the AuNP surface by the addition of high concentrations of 2-mercaptoethanol (2-ME), which binds strongly to the AuNPs. The sensitivity of this method to indoleamines could be significantly enhanced by increasing the AuNP concentration, incubation time, and sample volume. Under optimal extraction and separation conditions, the combination of NP-based extraction and CE-LINF provided 48-, 4077-, 985-, 920-, and 4030-fold improvements in the limits of detection (signal-to-noise ratio of 3) for 5-HTP, Trp, TA, 5-HT, and 5-HIAA as compared to the analysis of five indoleamines by CE-LINF. In addition, this proposed method was successfully used for the determination of TA and 5-HT in urine.     

Using nile red-adsorbed gold nanoparticles to locate glutathione within erythrocytes

An aqueous solution of Nile Red (NR)-absorbed 32-nm gold nanoparticles (AuNPs) have been used to sense glutathione (GSH). When the NR product is displaced by GSH on the AuNP surface, the fluorescence of the solution increases and the AuNPs aggregate. To determine the concentration and distribution of GSH within erythrocyte cells, a homemade fluorescence and scattering microscope was constructed. This system allows monitoring, within individual cells, of the uptake and transportation of the NRAuNPs and the displacement of the NR product from the NRAuNP surface by GSH. The fluorescence and scattering images clearly indicate the location of GSH inside the cells; these findings are supported by images recorded using 2,3-naphthalenedicarboxaldehyde, which is a highly selective fluorogenic reagent for GSH. Microscopic fluorescence measurements of the NRAuNPs revealed that the GSH concentration inside erythrocyte cells is 1.30 +/- 0.31 mM. To confirm this result, lysed erythrocyte cells were analyzed by applying capillary electrophoresis in conjunction with laser-induced fluorescence using NRAuNPs; accordingly, the average GSH concentration in a single erythrocyte cell was determined to be 1.32 +/- 0.06 mM.     

DNA analysis on microfabricated electrophoretic devices with bubble cells

Microfluidic devices with bubble cells have been fabricated on poly(methyl methacrylate) (PMMA) plates and have been employed for the analysis of DNA using polyethylene oxide (PEO) solutions. First, the separation channel was fabricated using a wire-imprinting method. Then, wires with greater sizes or a razor blade glued in a polycarbonate plate was used to fabricate bubble cells, with sizes of 190-650 microm. The improvements in resolution and sensitivity have been achieved for large DNA (> 603 base pair, bp) using such devices, which depend on the geometry of the bubble cell. The main contributor for optimal resolution is mainly due to DNA migration at lower electric field strengths inside the bubble cell. On the other hand, slight losses of resolution for small DNA fragments have been found mainly due to diffusion, supported by the loss of resolution when separating two small solutes. With a bubble cell of 75 microm (width) x 500 microm (depth), the sensitivity improvement up to 17-fold has been achieved for the 271 bp fragment in the separation of PhiX-174/HaeIII DNA restriction fragments. We have also found that a microfluidic device with a bubble cell of 360 microm x 360 microm is appropriate for DNA analysis. Such a device has been used for separating DNA ranging from 8 to 2176 bp and polymerase chain reaction (PCR) products amplified after 30 cycles, with rapidity and improvements in the sensitivity as well as resolution.     

Amplification of small analytes in polymer solution by capillary electrophoresis

We report concentration methods for the analysis of small solutes by capillary electrophoresis in conjunction with laser-induced native fluorescence using a Nd:YAG laser. After injecting samples, poly(ethylene oxide) (PEO) in the anode reservoir entered a capillary filled with Tris-borate buffer. When migrating in PEO solution, the analytes slowed down and stacked at the interface between the sample zone and PEO solution. As a result, the limits of detection (LODs) down to 8 pM for 2-naphthalenesulfonic acid and 70 pM for L-tryptophan have been achieved when injecting at 30 cm height for 120 s and 230 s, respectively. Such low LODs are partially due to the effects of NaCl in the samples and PEO on the fluorescence characteristics of the analytes. In addition, the concentrations of NaCl and PEO have great impacts on the migration of the analytes and electroosmotic flow, thereby affecting resolution and speed. Without pretreatment, the determinations of five important markers in urine samples and two acids in a cerebrospinal fluid sample have been performed separately, with the relative standard deviations of the concentrations less than 3.6%. Furthermore, by applying a short plug of low-pH buffer after injection, the analysis of greater volumes of the urine sample has been carried out, resulting in detecting more peaks.     

Incorporating peptides in the outer-coordination sphere of bioinspired electrocatalysts for hydrogen production

Four new cyclic 1,5-diaza-3,7-diphosphacyclooctane ligands have been prepared and used to synthesize [Ni(P(Ph)(2)N(R)(2))(2)](2+) complexes in which R is a mono- or dipeptide. These complexes represent a first step in the development of an outer-coordination sphere for this class of complexes that can mimic the outer-coordination sphere of the active sites of hydrogenase enzymes. Importantly, these complexes retain the electrocatalytic activity of the parent [Ni(P(Ph)(2)N(Ph)(2))(2)](2+) complex in an acetonitrile solution with turnover frequencies for hydrogen production ranging from 14 to 25 s(-1) in the presence of p-cyanoanilinium trifluoromethanesulfonate and from 135 to 1000 s(-1) in the presence of protonated dimethylformamide, with moderately low overpotentials, ～0.3 V. The addition of small amounts of water results in rate increases of 2-7 times. Unlike the parent complex, these complexes demonstrate dynamic structural transformations in solution. These results establish a building block from which larger peptide scaffolding can be added to allow the [Ni(P(R)(2)N(R')(2))(2)](2+) molecular catalytic core to begin to mimic the multifunctional outer-coordination sphere of enzymes.     

A short enantioselective synthesis of N-Boc-(2R,3R)-3-methyl-3-hydroxypipecolic acid from geraniol

The asymmetric synthesis of (2R,3R)-3-methyl-3-hydroxypipecolic acid, a key intermediate in the synthesis of dual MMP-13/aggrecanase inhibitors, is described. The title compound is prepared in seven steps with an overall yield of 41% starting from geraniol. Key steps in the synthesis include Sharpless asymmetric epoxidation, which establishes the chiral centers, and a one-pot oxidative olefin cleavage/reductive amination sequence that closes the piperidine ring.     

Colorimetric detection of lysozyme based on electrostatic interaction with human serum albumin-modified gold nanoparticles

In this study, an aqueous solution of 13-nm gold nanoparticles (AuNPs) covalently bonded with human serum albumin (HSA) was used for sensing lysozyme (Lys). HSA molecules were good stabilizing agents for AuNPs in high-salt solution and exhibited the ability to bond with Lys electrostatically. The aggregation of HSA-AuNPs was achieved upon the addition of high-pI proteins, such as Lys, alpha-chymotrypsinogen A, and conalbumin. Not the same was achieved, however, when low-pI proteins such as ovalbumin, bovine serum albumin, and alpha-lactalbumin were added. Matrix-assisted desorption/ionization mass spectrometry was used to demonstrate the interaction between HSA-AuNPs and Lys. It was found that the sensitivity of HSA-AuNPs for Lys was highly dependent on the HSA concentration. The Lys-induced aggregation of HSA-AuNPs was suggested based on the London-van der Waals attractive force. We further improved the selectivity of the probe by adjusting the pH solution to 8.0. Under the optimum conditions, the selectivity of this system for Lys over other proteins in high-salt solutions was remarkably high, even when their pI was very close to the Lys. The lowest detectable concentration of Lys in this approach was 50 nM. The applicability of the method was validated through the analyses of Lys in chicken egg white.     

Enrichment and separation of acidic and basic proteins using the centrifugal ultrafiltration followed by nanoparticle-filled capillary electrophoresis

This report describes a method for enrichment and separation of acidic and basic proteins using the centrifugal ultrafiltration followed by nanoparticle-filled capillary electrophoresis. To improve stacking and separation efficiencies of proteins, the separation buffer containing 1.6% poly(diallyldimethylammonium chloride) was added with gold nanoparticles (AuNP), poly(ethylene oxide), cetyltrimethylammonium bromide, and poly(vinyl alcohol). As a result, the use of AuNP as additives exhibited better efficiency in separation, stacking, and analysis time. Even for large-volume samples (110 nL), the separation efficiencies of acidic and basic proteins remained greater than 10(4) and 10(5) plates/m, respectively. To further enhance detection sensitivity, protein samples were enriched using the centrifugal ultrafiltration, followed by our proposed stacking method. The detection sensitivity was improved up to 314-fold compared to normal hydrodynamic injection. Additionally, the limits of detection at a signal-to-noise of 3 for most proteins were down to nanomolar range. We have validated the application of our method by means of analyses of 50 nM lysozyme in saliva samples. The proposed method was also successfully applied to the analyses of egg-white proteins, which have large differences in molecular weight and pI.