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The redox metabolic and bioenergetic paradigm of murburn concept advocates that diffusible reactive species (DRS, particularly oxygen-centric 1e-active radicals) are a crucial mainstay of physiology, and not mere pathological... more
The redox metabolic and bioenergetic paradigm of murburn concept advocates that diffusible reactive species (DRS, particularly oxygen-centric 1e-active radicals) are a crucial mainstay of physiology, and not mere pathological manifestations. The murburn purview of cellular function also integrates the essential principles of bioenergetics, thermogenesis, homeostasis, electrophysiology and coherence. In this context, any enzyme that generates/utilizes/sustains DRS functionality is called a murzyme. We have demonstrated that several water-soluble (hemoglobin, lactate dehydrogenase, peroxidases, etc.) and membrane-embedded (Complexes I-V in mitochondria, Photosystems I/II in chloroplasts, rhodopsin/transducin in rod cells, etc.) proteins serve as murzymes (Manoj & Gideon, BBA-Biomem., 2022). In the first part of our work on NKA (Na,K-ATPase), we had effectively questioned the classical mechanistic understanding of how this protein supposedly aids cells by stoichiometrically pumping 3 N...
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: Drosophila melanogaster is a prominent organism in developmental biology research and in studies related to pathophysiological conditions like cancer and Alzheimer’s disease. The fruit fly gut contains several cytochrome P450s... more
: Drosophila melanogaster is a prominent organism in developmental biology research and in studies related to pathophysiological conditions like cancer and Alzheimer’s disease. The fruit fly gut contains several cytochrome P450s (CYP450s), which have central roles in Drosophila development and in the normal physiology of the gut. Since the crystal structures of these proteins have not been deciphered yet, we modeled the structure of 29 different D. melanogaster gut CYP450s using Prime (Schrödinger). The sequences of chosen D. melanogaster gut CYP450s were compared with that of their human counterparts. The common gut (and liver) microsomal CYP450s in humans were chosen for structural comparison to find the homology and identity % of D. melanogaster CYPs with that of their human counterparts. The modeled structures were validated using PROCHECK and the best fit models were used for docking several known human pharmacological agents/drugs to the modeled D. melanogaster gut CYP450s. Based on the binding affinities (ΔG values) of the selected drug molecules with the modeled fly gut CYPs, the plausible differences in metabolism of the prominent drugs in humans and flies were projected. The gut is involved in the absorption of oral drugs/pharmacological agents, and hence, upregulation of intestinal CYP450 and their reactions with endobiotics and xenobiotics is envisaged. The insights gleaned from this work can validate D. melanogaster as a model organism for studying intestinal drug metabolism, particularly in the context of a) toxicology of pharmacological agents to the gut cells and b) how gut P450 metabolites/products can influence gut homeostasis. This work can help establish a platform for further in vitro investigations on how intestinal CYP450 metabolism can influence gut health. The data from this work can be used for further in silico studies and this work can serve as a platform for future in vitro investigations on intestinal CYP450-mediated metabolism of endo- and xeno-biotics in D. melanogaster.
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It is unresolved why lactate is transported to the liver for further utilization within the physiological purview of Cori cycle, when muscles have more lactate dehydrogenase (LDH) than liver. We point out that the answer lies in... more
It is unresolved why lactate is transported to the liver for further utilization within the physiological purview of Cori cycle, when muscles have more lactate dehydrogenase (LDH) than liver. We point out that the answer lies in thermodynamics/equilibriums. While the utilization of NADH for the reduction of pyruvate to lactate can be mediated via the classical mechanism, the oxidation of lactate (with/without the uphill reduction of NAD+) necessitates alternative physiological approaches. The latter pathway occurs via interactive equilibriums involving the enzyme, protons and oxygen or diffusible reactive oxygen species (DROS). Since liver has high DROS, the murburn activity at LDH would enable the cellular system to tide over the unfavorable energy barriers of the forward reaction (~476 kJ/mol; earlier miscalculated as ~26 kJ/mole). Further, the new mechanism does not necessitate any “smart decision‐making” or sophisticated control by/of proteins. The DROS‐based murburn theory explains the invariant active‐site structure of LDH isozymes and their multimeric nature. The theoretical insights, in silico evidence and analyses of literature herein also enrich our understanding of the underpinnings of “lactic acidosis” (lowering of physiological pH accompanied by lactate production), Warburg effect (increased lactate production at high pO2 by cancer cells) and approach for cancer therapy.
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Viruses, being obligate intracellular parasites, must first attach themselves and gain entry into host cells. Viral fusion machinery is the central player in the viral attachment process in almost every viral disease. Viruses have... more
Viruses, being obligate intracellular parasites, must first attach themselves and gain entry into host cells. Viral fusion machinery is the central player in the viral attachment process in almost every viral disease. Viruses have incorporated an array of efficient fusion proteins on their surfaces to bind efficiently to host cell receptors. They make use of the host proteolytic enzymes to rearrange their surface protein(s) into the form which facilitates their binding to host-cell membrane proteins and subsequently, fusion. This stage of viral entry is very critical and has many therapeutic implications. The current global pandemic of COVID-19 has sparked severe health crisis and economic shutdowns. SARS-CoV2, the etiological agent of the disease has led to millions of deaths and brought the scientific community together in an attempt to understand the mechanisms of SARS-CoV2 pathogenesis and mortality. Like other viral fusion machinery, CoV2 spike (S) glycoprotein- ‘The Demogorgon...
Research Interests: Biology and Cell Biology
The classical paradigm of visual physiology comprises of the following features: (i) rod/cone cells located at the rear end of the retina serve as the primary transducers of incoming photo-information, (ii) cis-trans retinal (C20H28O)... more
The classical paradigm of visual physiology comprises of the following features: (i) rod/cone cells located at the rear end of the retina serve as the primary transducers of incoming photo-information, (ii) cis-trans retinal (C20H28O) transformations on rhodopsin act as the transduction switch to generate a transmittable signal, (iii) signal amplification occurs via GDP-GTP exchange at transducin, and (iv) the amplified signal is relayed (as an action potential) as a flux-based ripple of Na-K ions along the axons of neurons. Fundamental physical principles, chemical kinetics and awareness of architecture of eye/retina prompt a question of these classical assumptions. In lieu, based on experimental and in silico findings, a simple space-time resolved murburn model for the physiology of photo-transduction in the retina is presented wherein molecular oxygen plays key roles. It is advocated that: (a) photo-induced oxygen to superoxide conversion serves as the key step in signal transduc...
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Isatin (1H-indole-2,3-dione)-containing compounds have been shown to possess several remarkable biological activities. We had previously explored a few isatin-based imidazole derivatives for their predicted dual activity against both... more
Isatin (1H-indole-2,3-dione)-containing compounds have been shown to possess several remarkable biological activities. We had previously explored a few isatin-based imidazole derivatives for their predicted dual activity against both inflammation and cancer. We explored 47 different isatin-based derivatives (IBDs) for other potential biological activities using <i>in silico</i> tools and found them to possess anti-viral activity. Using AutoDock tools, the binding site, binding energy, inhibitory constant/<i>K</i> <sub>i</sub> and receptor-ligand interactions for each of the compounds were analyzed against SARS-CoV-2 RNA-dependent RNA polymerase (RdRp). The partition coefficient (logP) values were predicted using MedChem Designer tool. Based on the best <i>K</i> <sub>i</sub>, binding energy and the ideal range of logP (between 1.0 and 3.0), 10 out of total 47 compounds were deemed to be prospective RdRp inhibitors. Some of t...
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We recently proposed a diffusible reactive (oxygen) species (DRS/DROS) based function for cytochrome b complexes (CBC) and quinones (Q)/quinols (QH2) in the murburn model of bioenergetics. This proposal is in direct conflict with the... more
We recently proposed a diffusible reactive (oxygen) species (DRS/DROS) based function for cytochrome b complexes (CBC) and quinones (Q)/quinols (QH2) in the murburn model of bioenergetics. This proposal is in direct conflict with the classical purview of Q‐cycle. Via extensive analyses of the structure‐function correlations of membrane‐quinones/quinols and proteins, we present qualitative and quantitative arguments to infer that the classical model cannot explain the energetics, kinetics, mechanism and probabilistic considerations. Therefore, it is proposed that Q‐cycle is neither necessary nor feasible at CBCs. In contrast, we substantiate that the murburn model explains: (a) crucial structural data of CBCs, (b) why quinones/quinols are utilized in bioenergetic membranes, (c) how trans‐membrane potential is generated owing to effective charge separation at CBCs, (d) mobility data of O2, DRS, Q/QH2, and (e) utility of other reaction/membrane components. Further, the murburn model also accommodates the absence of quinones in anaerobic Archaea, wherein methanophenazines are prevalent. The work mandates that the textbooks and research agendas are refreshed to reflect the new perception.
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Mitochondrial membrane‐embedded redox proteins are classically perceived as deterministic “electron transport chain” (ETC) arrays cum proton pumps; and oxygen is seen as an “immobile terminal electron acceptor.” This is untenable because:... more
Mitochondrial membrane‐embedded redox proteins are classically perceived as deterministic “electron transport chain” (ETC) arrays cum proton pumps; and oxygen is seen as an “immobile terminal electron acceptor.” This is untenable because: (1) there are little free protons to be pumped out of the matrix; (2) proton pumping would be highly endergonic; (3) ETC‐chemiosmosis‐rotary ATP synthesis proposal is “irreducibly complex”/”non‐evolvable” and does not fit with mitochondrial architecture or structural/distribution data of the concerned proteins/components; (4) a plethora of experimental observations do not conform to the postulates/requisites; for example, there is little evidence for viable proton‐pumps/pH‐gradient in mitochondria, trans‐membrane potential (TMP) is non‐fluctuating/non‐trappable, oxygen is seen to give copious “diffusible reactive (oxygen) species” (DRS/DROS) in milieu, etc. Quite contrarily, the newly proposed murburn model's tenets agree with known principles of energetics/kinetics, and builds on established structural data and reported observations. In this purview, oxygen is needed to make DRS, the principal component of mitochondrial function. Complex V and porins respectively serve as proton‐inlet and turgor‐based water‐exodus portals, thereby achieving organellar homeostasis. Complexes I to IV possess ADP‐binding sites and their redox‐centers react/interact with O2/DRS. At/around these complexes, DRS cross‐react or activate/oxidize ADP/Pi via fast thermogenic one‐electron reaction(s), condensing to form two‐electron stabilized products (H2O2/H2O/ATP). The varied architecture and distribution of components in mitochondria validate DRS as (i) the coupling agent of oxidative reactions and phosphorylations, and (ii) the primary reason for manifestation of TMP in steady‐state. Explorations along the new precepts stand to provide greater insights on mitochondrial function and pathophysiology.
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Thermogenesis by uncoupling protein (UCP) has traditionally been explained as the dissipation of proton gradient across the inner mitochondrial membrane into heat. Herein, we propose that UCPs, aided by the large pore and positively... more
Thermogenesis by uncoupling protein (UCP) has traditionally been explained as the dissipation of proton gradient across the inner mitochondrial membrane into heat. Herein, we propose that UCPs, aided by the large pore and positively charged amino acids of suspended loops, enable protonation and transport of DROS. Thus, UCP facilitates DROS-reactions amongst themselves, forming water and liberating heat around the inner mitochondrial membrane. Thereby, the simple murburn scheme for biothermogenesis integrates structural information of UCP with its attributed physiological function.
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In this second half of our treatise on oxygenic photosynthesis, we provide support for the murburn model of the light reaction of photosynthesis and ratify key predictions made in the first part. Molecular docking and visualization of... more
In this second half of our treatise on oxygenic photosynthesis, we provide support for the murburn model of the light reaction of photosynthesis and ratify key predictions made in the first part. Molecular docking and visualization of various ligands of quinones/quinols (and their derivatives) with PS II/Cytochrome b6f complexes did not support chartered 2e-transport role of quinols. A broad variety of herbicides did not show any affinity/binding-based rationales for inhibition of photosynthesis. We substantiate the proposal that disubstituted phenolics (perceived as protonophores/uncouplers or affinity-based inhibitors in the classical purview) serve as interfacial modulators of diffusible reactive (oxygen) species or DR(O)S. The DRS-based murburn model is evidenced by the identification of multiple ADP-binding sites on the extra-membraneous projection of protein complexes and structure/distribution of the photo/redox catalysts. With a panoramic comparison of the redox metabolic machinery across diverse organellar/cellular systems, we highlight the ubiquitous one-electron murburn facets (cofactors of porphyrin, flavin, FeS, other metal centers and photo/redox active pigments) that enable a facile harnessing of the utility of DRS. In the summative analyses, it is demonstrated that the murburn model of light reaction explains the structures of membrane supercomplexes recently observed in thylakoids and also accounts for several photodynamic experimental observations and evolutionary considerations. In toto, the work provides a new orientation and impetus to photosynthesis research.Communicated by Ramaswamy H. Sarma.
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Isatin (1H-indole-2,3-dione)-containing compounds have been shown to possess several remarkable biological activities. We had previously explored a few isatin-based imidazole derivatives for their predicted dual activity against both... more
Isatin (1H-indole-2,3-dione)-containing compounds have been shown to possess several remarkable biological activities. We had previously explored a few isatin-based imidazole derivatives for their predicted dual activity against both inflammation and cancer. We explored 47 different isatin-based derivatives (IBDs) for other potential biological activities using in silico tools and found them to possess anti-viral activity. Using AUTODOCK tools, the binding site, binding energy, inhibitory constant/Ki and receptor-ligand interactions for each of the compounds was analyzed against SARS-CoV-2 RNA-dependent RNA polymerase (RdRp). The partition coefficient (logP) values were predicted using MedChem Designer tool. Based on the best Ki, binding energy and the ideal range of logP (between 1.0-3.0), 10 out of total 47 compounds were deemed to be prospective RdRp inhibitors. Some of these compounds gave better Ki, binding energy and logP values when compared to standard RdRp inhibitors such a...
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Quinones are found in the lipid-membranes of prokaryotes like E. coli and cyanobacteria, and are also abundant in eukaryotic mitochondria and chloroplasts. They are intricately involved in the reaction mechanism of redox phosphorylations.... more
Quinones are found in the lipid-membranes of prokaryotes like E. coli and cyanobacteria, and are also abundant in eukaryotic mitochondria and chloroplasts. They are intricately involved in the reaction mechanism of redox phosphorylations. In the Mitchellian chemiosmotic school of thought, membrane-lodged quinones are perceived as highly mobile conveyors of two-electron equivalents from the first leg of Electron Transport Chain (ETC) to the ‘second pit-stop’ of Cytochrome bc1 or b6f complex (CBC), where they undergo a regenerative ‘Q-cycle’. In Manoj’s murburn mechanism, the membrane-lodged quinones are perceived as one- or two- electron donors/acceptors, enabling charge separation and the CBC resets a one-electron paradigm via ‘turbo logic’. Herein, we compare various purviews of the two mechanistic schools with respect to: constraints in mobility, protons’ availability, binding of quinones with proteins, structural features of the protein complexes, energetics of reaction, overall ...
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One of the most fundamental questions in biology pertains to how mechano-chemical energy is derived from metabolic fuels. In particular, how oxidation of NADH is linked to ATP synthesis in mitochondrial oxidative phosphorylation (mOxPhos)... more
One of the most fundamental questions in biology pertains to how mechano-chemical energy is derived from metabolic fuels. In particular, how oxidation of NADH is linked to ATP synthesis in mitochondrial oxidative phosphorylation (mOxPhos) has been a topic of intense debate. Together, the Peter Mitchell-Paul Boyer proposals for mOxPhos are termed herein as “chemiosmotic rotary ATP synthesis” (or CRAS) model, which was recently defended/advocated by Pedro Silva in Biophysical Chemistry . Over the last two decades, Sunil Nath had questioned some aspects of the CRAS proposal, and made subtle alterations on the roles of Complex V and ions within the reaction scheme, and continues to advocate his framework as “two-ion torsional ATP synthesis” (abbreviated herein as TITAS) model in Biophysical Chemistry . Kelath Murali Manoj had revisited the data on the respiratory machinery’s structures/distributions and based on two-decades of evidence-based experimental research in redox enzymology of ...
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We summarize salient components of the Z-Scheme electron transport chain within chloroplasts. Banking on the skepticism kindled by Robert Emerson’s reproducible observations, we explore the working of its various elements using updated... more
We summarize salient components of the Z-Scheme electron transport chain within chloroplasts. Banking on the skepticism kindled by Robert Emerson’s reproducible observations, we explore the working of its various elements using updated information on the distribution of components within the chloroplast architecture, structure-function correlations of relevant redox proteins, thermodynamics, kinetics, and evolutionary principles. Through a set of simple models/simulations and a series of intriguing queries, we moot several theoretical premises and highlight evidences/arguments that question the physiological feasibility of Z-scheme. Finally, we also point to new avenues for furthering research in this field.
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The inefficiency of cyanide/HCN (CN) binding with heme proteins (under physiological regimes) is demonstrated with an assessment of thermodynamics, kinetics, and inhibition constants. The acute onset of toxicity and CN’s mg/Kg LD50(μM... more
The inefficiency of cyanide/HCN (CN) binding with heme proteins (under physiological regimes) is demonstrated with an assessment of thermodynamics, kinetics, and inhibition constants. The acute onset of toxicity and CN’s mg/Kg LD50(μM lethal concentration) suggests that the classical hemeFe binding-based inhibition rationale is untenable to account for the toxicity of CN. In vitro mechanistic probing of CN-mediated inhibition of hemeFe reductionist systems was explored as a murburn model for mitochondrial oxidative phosphorylation (mOxPhos). The effect of CN in haloperoxidase catalyzed chlorine moiety transfer to small organics was considered as an analogous probe for phosphate group transfer in mOxPhos. Similarly, inclusion of CN in peroxidase-catalase mediated one-electron oxidation of small organics was used to explore electron transfer outcomes in mOxPhos, leading to water formation. The free energy correlations from a Hammett study and IC50/Hill slopes analyses and comparison w...
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Recently, electron transfers and catalyses in a bevy of redox reactions mediated by hemeproteins were explained by murburn concept. The term "murburn" is abstracted from "" or "" and connotes a novel... more
Recently, electron transfers and catalyses in a bevy of redox reactions mediated by hemeproteins were explained by murburn concept. The term "murburn" is abstracted from "" or "" and connotes a novel "" interaction paradigm. Quite unlike the genetic regulations and protein-level affinity-based controls that govern order and specificity/selectivity in conventional treatments, murburn concept is based on stochastic/thermodynamic regulatory principles. The novel insight necessitates a "reactivity outside the active-site" perspective, because select redox enzymatic activity is obligatorily mediated via diffusible radical/species. Herein, reactions employing key hemeproteins (as exemplified by CYP2E1) establish direct experimental connection between "additive-influenced redox catalysis" and "unusual dose responses" in reductionist and physiological milieu. Thus, direct and conclusive molecular-level experimental eviden...
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Peroxidations mediated by heme-enzymes have been traditionally studied under a single-site (heme distal pocket), non-sequential (ping-pong), two-substrates binding scheme of Michaelis-Menten paradigm. We had reported unusual modulations... more
Peroxidations mediated by heme-enzymes have been traditionally studied under a single-site (heme distal pocket), non-sequential (ping-pong), two-substrates binding scheme of Michaelis-Menten paradigm. We had reported unusual modulations of peroxidase and P450 reaction outcomes and explained it invoking diffusible reactive species [Manoj, 2006; Manoj et al., 2010; Andrew et al., 2011, Parashar et al., 2014 & Venkatachalam et al., 2016]. A systematic investigation of specific product formation rates was undertaken to probe the hypothesis that involvement of diffusible reactive species could explain undefined substrate specificities and maverick modulations (sponsored by additives) of heme-enzymes. When the rate of specific product formation was studied as a function of reactants' concentration or environmental conditions, we noted marked deviations from normal profiles. We report that heme-enzyme mediated peroxidations of various substrates are inhibited (or activated) by sub-equi...
Research Interests: Biochemistry, Chemistry, Catalysis, Drug metabolism, Cytochrome C, and 15 moreCytochrome c oxidase, Catalase, Heme proteins, Heme, Enzyme, Biochimie, Active site, Binding, Allosteric modulators, Hormesis, Ascomycota, Electronic Transport In Molecules, Biochemistry and cell biology, Biological oxidation, and chloroperoxidase
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Within the context of light reaction of photosynthesis, the structure-function correlations of the chloroplast proteins of plastocyanin and ferredoxins (Fd) are analyzed via two perspectives: 1) The Z-scheme, which considers PC/Fd as... more
Within the context of light reaction of photosynthesis, the structure-function correlations of the chloroplast proteins of plastocyanin and ferredoxins (Fd) are analyzed via two perspectives: 1) The Z-scheme, which considers PC/Fd as specific affinity binding-based electron-relay agents, thereby deterministically linking the functions of Cytochrome b6f (Cyt. b6f) and Photosystem I (PS I) to NADP+ reduction by Fd:NADPH oxidoreductase (FNR) via protein-protein contacts and 2) The murburn explanation for oxygenic photophosphorylation, which deems PC/Fd as generic ‘redox capacitors’, temporally accepting and releasing one-electron equivalents in reaction milieu. Amino acid residues located on the surface loci of key patches of PC/Fd vary in electrostatic/contour (topography) signatures. Crystal structures of four different complexes each of cyt.f-PC and Fd-FNR show little conservation in the contact-surfaces, thereby discrediting ‘affinity binding-based electron transfers (ET)’ as an ev...
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Hitherto, electron transfer (ET) between redox proteins has been deemed to occur via donor–acceptor binding, and diffusible reactive species are considered as deleterious side-products in such systems. Herein, ET from cytochrome P450... more
Hitherto, electron transfer (ET) between redox proteins has been deemed to occur via donor–acceptor binding, and diffusible reactive species are considered as deleterious side-products in such systems. Herein, ET from cytochrome P450 reductase (CPR, an animal membrane flavoprotein) and horseradish peroxidase (HRP, a plant hemoprotein) to cytochrome c (Cyt c, a soluble animal hemoprotein) was probed under diverse conditions, using standard assays. ET in the CPR-Cyt c system was critically inhibited by cyanide and sub-equivalent levels of polar one-electron cyclers like copper ions, vitamin C/ Trolox and superoxide dismutase. In the presence of lipids, inhibition was also afforded by amphipathic molecules vitamin E, palmitoyl-vitamin C and the membrane hemoprotein, cytochrome b 5. Such non-specific inhibition (by diverse agents in both aqueous and lipid phases) indicated that electron transfer/ relay was effected by small diffusible agents, whose lifetimes are shortened by the diverse radical scavengers. When CPR was retained in a dialysis membrane and Cyt c presented outside in free solution, ET was still observed. Further, HRP (taken at nM levels) catalyzed oxidation of a phenolic substrate was significantly inhibited upon the incorporation of sub-nM levels of Cyt c. The findings imply that CPR-Cyt c or HRP-Cyt c binding is not crucial for ET. Further, fundamental quantitative arguments (based on diffusion/collision) challenge the erstwhile protein–protein binding-assisted ET hypothesis. It is proven beyond reasonable doubt that mobile and diffusible electron carriers (ions and radicals) serve as " redox-relay agents " in the biological ET models/setup studied.