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1.
The binding affinity of the two substrate–water molecules to the water-oxidizing Mn4CaO5 catalyst in photosystem II core complexes of the extremophilic red alga Cyanidioschyzon merolae was studied in the S2 and S3 states by the exchange of bound 16O-substrate against 18O-labeled water. The rate of this exchange was detected via the membrane-inlet mass spectrometric analysis of flash-induced oxygen evolution. For both redox states a fast and slow phase of water-exchange was resolved at the mixed labeled m/z 34 mass peak: kf = 52 ± 8 s− 1 and ks = 1.9 ± 0.3 s− 1 in the S2 state, and kf = 42 ± 2 s− 1 and kslow = 1.2 ± 0.3 s− 1 in S3, respectively. Overall these exchange rates are similar to those observed previously with preparations of other organisms. The most remarkable finding is a significantly slower exchange at the fast substrate–water site in the S2 state, which confirms beyond doubt that both substrate–water molecules are already bound in the S2 state. This leads to a very small change of the affinity for both the fast and the slowly exchanging substrates during the S2 → S3 transition. Implications for recent models for water-oxidation are briefly discussed.  相似文献   

2.
Human M-proinsulin was cleaved by trypsin at the R31R32–E33 and K64R65–G66 bonds (B/C and C/A junctions), showing the same cleavage specificity as exhibited by prohormone convertases 1 and 2 respectively. Buffalo/bovine M-proinsulin was also cleaved by trypsin at the K59R60–G61 bond but at the B/C junction cleavage occurred at the R31R32–E33 as well as the R31–R32E33 bond. Thus, the human isoform in the native state, with a 31 residue connecting C-peptide, seems to have a unique structure around the B/C and C/A junctions and cleavage at these sites is predominantly governed by the structure of the proinsulin itself. In the case of both the proinsulin species the cleavage at the B/C junction was preferred (65%) over that at the C/A junction (35%) supporting the earlier suggestion of the presence of some form of secondary structure at the C/A junction. Proinsulin and its derivatives, as natural substrates for trypsin, were used and mass spectrometric analysis showed that the kcat./Km values for the cleavage were most favourable for the scission of the bonds at the two junctions (1.02 ± 0.08 × 105 s− 1 M− 1) and the cleavage of the K29–T30 bond of M-insulin-RR (1.3 ± 0.07 × 105 s− 1 M− 1). However, the K29–T30 bond in M-insulin, insulin as well as M-proinsulin was shielded from attack by trypsin (kcat./Km values around 1000 s− 1 M− 1). Hence, as the biosynthetic path follows the sequence; proinsulin → insulin-RR → insulin, the K29–T30 bond becomes shielded, exposed then shielded again respectively.  相似文献   

3.
The driver tries to keep the car in the center of the lane. If the car is too near the left edge, this causes the driver to make a “corrective” right turn. If the car is near the right edge, a “corrective” left turn is made. Therefore, a quantity which decreases with increasing distance Δ L from the left edge may be considered as a stimulusS R producing the reactionR R of turning to the right. A similar situation holds for the distance Δ R from the right edge. When the car is in the center of the lane, Δ L = Δ R andS R =S L , the stimuli are equal. We thus have here a situation analogous to the one studied by H. D. Landahl in his theory of psychophysical discrimination. In general a reactionR R (resp.R L ) will occur only ifR R R L h * (resp.R L R R h *) whereh * is a threshold. Applying Landahl’s theory to this situation, we find thath * determines the distance from the edge, at which a corrective turn is made. This distance is not constant, but a function of the speedv of the car. The requirement that a corrective turn should be madebeforre the car runs off the road leads to an expression for the maximum safe speed. Because of the transcendency of the equations involved, closed solutions cannot be obtained. It is, however, shown that the expression for maximum safe speed, given in a previous paper (Bull. Math. Biophysics,21, 299–308, 1959), is a rough first approximation to the expressions found now.  相似文献   

4.
The lipid dynamics of the adrenocortical microsomal membranes was studied by monitoring the fluorescence anisotropy and excited state lifetime of a set of anthroyloxy fatty acid probes (2-, 7-, 9- and 12-(9-anthroyloxy)-stearic acid (AP) and 16-(9-anthroyloxy)palmitic acid (AS). It was found that a decreasing polarity gradient from the aqueous membrane interface to the membrane interior, was present. This gradient was not modified by the proteins, as evidenced by comparison of complete membranes and derived liposomes, suggesting that the anthroyloxy probes were not in close contact with the proteins. An important change of the value of the mean rotational relaxation time as a function of the position of the anthroyl ring along the acyl chain was evidenced. In the complete membranes, a relatively more fluid medium was evidenced in the C16 as compared to the C2 region, while the rotational motion appeared to be the most hindered at the C7–C9 level. In the derived liposomes, a similar trend was observed but the mobility was higher at all levels. The decrease of the mean rotational relaxation time was more important for 12-AS and 16-AP. Temperature dependence of the mean rotational relaxation time of 2-AS, 12-AS and 16-AP in the complete membranes revealed the existence of a lipid reorganization occurring around 27°C and concerning mainly the C16 region. The extent to which the acyl chain reacted to this perturbation at the C12 level depended on pH. The presence of proteins increased the apparent magnitude of this reorganization and also modified the critical temperature from approx. 23°C in the derived liposomes to approx. 27°C in the complete membranes. Thermal dependence of the maximum velocity of the 3-oxosteroid Δ54-isomerase, the second enzyme in the enzymatic sequence, responsible for the biosynthesis of the 3-oxo4-steroids in the adrenal cortex microsomes, was studied. The activation energy of the catalyzed reaction was found to be low and constant (2–5 kcal · mol?1) in the temperature range 16–40°C at pH 7.5, 8.5 and 9, corresponding to the minimum, intermediate and maximum rate, respectively. A drastic increase of the activation energy (20 kcal · mol?1) was observed at temperature below 16°C at pH 7.5. A correlated change of the pKESapp as function of temperature was detected; at 36°C pKESapp = 8.3 while at 13°C the value shifted to 8.7. The pH range of the group ionization was narrower at 13°C. In contrast with the behaviour of the 3β-hydroxy5-steroid dehydrogenase, the 3-oxosteroid Δ54-isomerase was apparently unaffected by the lipid reorganization at 27°C. It is suggested that this enzyme possesses a different and more fluid lipid environment than the bulk lipids.  相似文献   

5.
Chitosan (Ch) was chemically modified with ethylenesulfide (Es) under solvent-free conditions to give (ChEs), displaying a high content of thiol groups due to opening of the three member cyclic reagent. Elemental analysis showed a decrease in nitrogen content. This result indicated the incorporation of two ethylenesulfide molecules for each unit of the polymeric structure of the precursor biopolymer. Infrared spectroscopy, thermogravimetry, and 13C NMR in the solid state demonstrated the effectiveness of the reaction, with signals at 30 ppm for ChEs due to the change in the methylene group environment. Divalent metal uptake by chemically modified biopolymer gave the order Cu > Ni > Co > Zn, reflecting the corresponding acidity of these cations in bonding to the sulfur and the basic nitrogen atoms available on the pendant chains. The equilibrium data were fitted to Freundlich, Temkin, and Langmuir models. The maximum monolayer adsorption capacity for the cations was found to be 1.54 ± 0.02, 1.25 ± 0.03, 1.13 ± 0.01, and 0.83 ± 0.03 mmol g−1, respectively. The Langmuir model best explained the cation–sulfur bond interactions at the solid–liquid interface. The thermodynamics for these interactions gave exothermic enthalpic values of −43.02 ± 0.03, −28.72 ± 0.02, −26.27 ± 0.04, and −17.32 ± 0.02 kJ mol−1, respectively. The spontaneity of the systems is given by negative Gibbs free energies of −31.2 ± 0.1, −32.7 ± 0.1, −31.7 ± 0.1, and −32.2 ± 0.1 kJ mol−1, respectively, in spite of the unfavorable negative entropic values of −39 ± 1, −13 ± 1, −18 ± 1, and −49 ± 1 J K−1 mol−1 due to solvent ordering in the course of complexation. This newly synthesized biopolymer is presented as a chemically useful material for cation removal from aqueous solution.  相似文献   

6.
The dissociation kinetics of the europium(III) complex with H8dotp ligand was studied by means of molecular absorption spectroscopy in UV region at ionic strength 3.0 mol dm−3 (Na,H)ClO4 and in temperature region 25-60 °C. Time-resolved laser-induced fluorescence spectroscopy (TRLIFS) was employed in order to determine the number of water molecules in the first coordination sphere of the europium(III) reaction intermediates and the final products. This technique was also utilized to deduce the composition of reaction intermediates in course of dissociation reaction simultaneously with calculation of rate constants and it demonstrates the elucidation of intimate reaction mechanism. The thermodynamic parameters for the formation of kinetic intermediate (ΔH0 = 11 ± 3 kJ mol−1, ΔS0 = 41 ± 11 J K−1 mol−1) and the activation parameters (Ea = 69 ± 8 kJ mol−1, ΔH = 67 ± 8 kJ mol−1, ΔS = −83 ± 24 J K−1 mol−1) for the rate-determining step describing the complex dissociation were determined. The mechanism of proton-assisted reaction was proposed on the basis of the experimental data.  相似文献   

7.
8.
Cytochrome c oxidase of the ba 3-type from Thermus thermophilus does not interact with cyanide in the oxidized state and acquires the ability to bind heme iron ligands only upon reduction. Cyanide complexes of the reduced heme a 3 in cytochrome ba 3 and in mitochondrial aa 3-type cytochrome oxidase are similar spectroscopically, but the a 32+-CN complex of cytochrome ba 3 is strikingly tight. Experiments have shown that the K d value of the cytochrome ba 3 complex with cyanide in the presence of reductants of the enzyme binuclear center does not exceed 10−8 M, which is four to five orders of magnitude less than the K d of the cyanide complex of the reduced heme a 3 of mitochondrial cytochrome oxidase. The tightness of the cytochrome ba 3 complex with cyanide is mainly associated with an extremely slow rate of the ligand dissociation (k off ≤ 10−7 sec−1), while the rate of binding (k on ∼ 102 M−1·sec−1) is similar to the rate observed for the mitochondrial cytochrome oxidase. It is proposed that cyanide dissociation from the cytochrome ba 3 binuclear center might be hindered sterically by the presence of the second ligand molecule in the coordination sphere of CuB2+. The rate of cyanide binding with the reduced heme a 3 does not depend on pH in the neutral area, but it approaches linear dependence on H+ activity in the alkaline region. Cyanide binding appears to be controlled by protonation of an enzyme group with pK a = 8.75.  相似文献   

9.
From the data of experiments with bees in which threshold response is employed as a means of recognizing visual discrimination between stripes of equal width alternately illuminated by intensities I 1 and I 2, it is shown that the detectable increment of intensity ΔI, where ΔI = I 2 - I 1, is directly proportional to σI2 (I 1 being fixed). From tests of visual acuity, where I 1 = 0 and the width of the stripes is varied, σI2 = kI 2 + const.; here I 2 = ΔI, and ΔI/I 2 = 1. When the visual excitability of the bee is changed by dark adaptation, λIkΔI (= k'' σΔI) = k'''' I + const. For the measurements of critical illumination at threshold response to flicker, σI2 (= σΔI) = k I 2 = k'' ΔI + const. The data for critical illumination producing threshold response to flicker in the sun-fish Lepomis show for the rods σI2 = K I 2 for the cones σI2 = K''(I 2 + const.). The data thus indicate that in all these experiments essentially the same visual function is being examined, and that the recognition of the production of a difference in effect by alternately illuminated stripes takes place in such a way that dI)/dI2) = const., and that ΔI is directly proportional to I (or "I 2," depending on the nature of the experiment). It is pointed out that the curve for each of the cases considered can be gotten equally well if mean I or σI is plotted as a function of the independent variable involved in the experiment. Certain consequences of these and related facts are important for the treatment of the general problem of intensity discrimination.  相似文献   

10.
We conducted a molecular analysis of the Vrn-B1 gene in two near-isogenic lines (NILs) carrying the dominant Vrn-B1 S and Vrn-B1 Dm alleles from the Saratovskaya 29 and Diamant 2 cultivars, respectively. These lines are characterized by different times of ear emergence. PCR analysis and subsequent sequencing of the regulatory regions of Vrn-B1 revealed the full identity of the promoter region in both alleles. Simultaneously, we found significant differences in the structure of the first intron of the Vrn-B1 S allele when compared to Vrn-B1 Dm ; specifically, the deletion of 0.8 kb coupled with the duplication of 0.4 kb. We suggest that these changes in intron 1 of Vrn-B1 S caused earlier ear emergence in the corresponding NIL. The unusual structure of intron 1 within the Vrn-B1 S allele was described for the first time in this study. The allele Vrn-B1 Dm was almost identical with the previously studied sequence of the Vrn-B1a allele of T. aestivum, Triple Dirk B. We designated the new Vrn-B1 S allele as Vrn-B1c. PCR analysis of the Vrn-B1 gene in 26 spring wheat cultivars of both Russian and foreign breeding revealed that 16 of them contain the Vrn-B1a allele and 6 contain the Vrn-B1c allele. Other cultivars studied contained the recessive vrn-B1 gene, except for Novosibirskaya 67. This study demonstrates that the traditional system of Vrn-1 markers does not fully encompass the allelic diversity of these genes because none of the cultivars containing the Vrn-B1c allele gave a PCR product using the previously developed set of primers for identification of the Vrn-B1 locus. We showed that the newly characterized Vrn-B1c allele is widely distributed among different genotypes of spring wheat. The findings indicate the impact of structural changes in the first intron of Vrn-1 on the vernalization response and heading time.  相似文献   

11.
Hydrochlorothiazide (HCTZ) was shown to inhibit the transepithelial NaCl transport and the apical Na+-Cl? symport and to depolarize the apical membrane potential in the rabbit gallbladder epithelium. The depolarization was likely related to the opening of a Cl? conductance. To better understand whether an apical Cl? leak is involved in the mechanism of action of HCTZ, the transapical Cl? backflux was measured radiochemically by the washout technique. The gallbladder wall, pretreated with pronase on the serosal side to homogenize the subepithelium, was loaded with 36Cl? on the luminal side; mucosal and serosal 36Cl? effluxes (J m , J s ) were then measured every 2 min. The pretreatment with pronase did not alter the membrane potentials and the selectivity of the epithelium. Under control conditions and the tissue in steady-state, J m and J s time courses were each described by two exponential decays (A,B); the rate constants, k A and k B , were 0.71 ±0.03 and 0.16±0.01 min?1, respectively, and correspondingly the half-times (t 1 2A , t 1 2B ) were 1.01±0.05 and 5.00±0.44 min (n=10); these parameters were not significantly different for J m and J s time courses. J s was always greater than J m (J s /J m =2.02±0.22 and 1.43 ±0.17 for A and B decays). Under SCN? treatment in steady-state conditions, both J m and J s time courses were described by only one exponential decay, the component B being abolished. Moreover t 1 2A was similar to that predictable for the subepithelium. It follows that it is the component B which exits the epithelial compartment. Based on the intracellular specific activity and 36Cl? J m B at 0 min time of the washout experiment, the cell-lumen Cl? backflux in steady-state was calculated to be equal to about 2 μmol cm?2hr?1, in agreement with the value indirectly computable by other techniques. The experimental model was well responsive to different external challenges (increases in media osmolalities; luminal treatment with nystatin). HCTZ (2.5 · 10?4 m) largely increased 36Cl? J m B . The increase was abolished by luminal treatment with 10?4 m SITS, which not only brought back the efflux time courses to the ones observed under control conditions but even increased J s /J m of the cellular component, an indication of a reduced J m B . It is concluded that HCTZ opens an apical, SITS-sensitive Cl? leak, which contributes to dissipate the intracellular Cl? accumulation and to inhibit the NaCl transepithelial transport. Moreover, the drug is likely to reduce the basal electroneutral Cl? backflux supported by Na+-Cl? cotransport, in agreement with the inhibition of the cotransport itself.  相似文献   

12.
13.
The effect of the protein structure of (Na+ + K+)-ATPase on its incorporation into liposome membranes was investigated as follows: the catalytic α-subunit of (Na+ + K+)-ATPase was split into low-molecular weight fragments by trypsin treatment and the digested enzyme was reconstituted at the same protein concentration as intact control enzyme. The reconstitution process was quantified by the average number of intramembrane particles appearing on concave and convex fracture faces after freeze-fracture of the (Na+ + K+)-ATPase liposomes. The number of intramembrane particles as well as their distribution on concave and convex fracture faces is not modified by the proteolysis. In contrast, the ATPase activity and the transport capacity of the (Na+ + K+)-ATPase decrease progessively with increasing incubation times in the presence of trypsin and are abolished when the original 100 000 molecular weight α-subunit is no longer visible by sodium dodecylsulfate gel electrophoresis. Apparently, functional (Na+ + K+)-ATPase with intact protein structure and digested, non functional enzyme consisting of fragments of the α-subunit reconstitute in the same manner and to the same extent as judged by freeze-fracture analysis. We conclude that, while trypsin treatment modifies the (Na+ + K+)-ATPase molecule in a functional sense, it appears not to modify its interaction with the bilayer in producing intramembrane particles. On the basis of our results, we propose a lipid-lipid interaction mechanism for reconstitution of (Na+ + K+)-ATPase.  相似文献   

14.
Abstract Three kinds of trisaccharides were prepared by digesting fucoidan from the brown alga Kjellmaniella crassifolia, with the extracellular enzymes of the marine bacterium Fucobacter marina. Their structures were determined as Δ4,5GlcpUA1-2(L-Fucp(3-O-sulfate)α1-3)D-Manp, Δ4,5GlcpUA1-2(L-Fucp(3-O-sulfate)α1-3)D-Manp(6-O-sulfate), and Δ4,5GlcpUA1-2(L-Fucp(2,4-O-disulfate)α1-3)D-Manp(6-O-sulfate), which indicated the existence of a novel polysaccharide in the fucoidan and a novel glycosidase in the extracellular enzymes. In order to determine the complete structure of the polysaccharide and the reaction mechanism of the glycosidase, the fucoidan was partially hydrolyzed to obtain glucuronomannan, which is the putative backbone of the polysaccharide, and its sugar sequence was determined as (-4-D-GlcpUAβ1-2D-Manpα1-)n, which disclosed that the main structure of the polysaccharide is (-4-D-GlcpUAβ1-2(L-Fucp(3-O-sulfate)α1-3)D-Manpα1-)n. Consequently, the glycosidase was deduced to be an endo-α-D-mannosidase that eliminatively cleaves the α-D-mannosyl linkage between D-Manp and D-GlcpUA residues in the polysaccharide and produces the above trisaccharides. The novel polysaccharide and glycosidase were tentatively named as sulfated fucoglucuronomannan (SFGM) and SFGM lyase, respectively.  相似文献   

15.
Previous studies have shown that the infectivity of baculovirus to herbivores is affected by phytochemicals ingested during the acquisition of viral inoculum on the foliage of host plants. Here, we measured the effects of 14 host plant species on the infectivity of Spodoptera exigua nucleopolyhedrovirus (SeNPV) to its larvae. The order of the LD50 values of SeNPV among the host plants was Ipomoea aquatica > Brassica oleracea > Raphanus sativus > Amaranthus tricolor > Spinacia oleracea > Vigna unguiculata > Solanum melongena > Capsicum annuum > Apium graveolens > Allium fistulosum > Lactuca sativa > Brassica chinensis > Zea mays > Glycine max, with 940.1 ± 2.26, 424.0 ± 0.60, 295.2 ± 1.13, 147.3 ± 0.63, 138.6 ± 0.22, 119.9 ± 0.07, 119.8 ± 0.02, 109.2 ± 0.18, 104.8 ± 0.62, 102.1 ± 0.66, 97.9 ± 0.22, 89.9 ± 0.32, 79.0 ± 0.13 and 64.0 ± 0.38 OBs per larva, respectively, and the values of mean time to death of virus‐infected larvae were 6.21 ± 0.11, 7.12 ± 0.10, 7.33 ± 0.21, 6.97 ± 0.02, 7.06 ± 0.01, 7.29 ± 0.03, 7.32 ± 0.05, 7.07 ± 0.08, 7.24 ± 0.11, 7.09 ± 0.13, 7.50 ± 0.06, 7.23 ± 0.01, 7.30 ± 0.02 and 7.19 ± 0.07 days, respectively. The mean time to death of larvae decreased with increasing viral dose, and corrected mortality decreased as the larval mean time to death increased. These findings have significance for understanding the effects of host plants on the infectivity of baculovirus to noctuids.  相似文献   

16.
17.
We explored the relationships between surface-soil (1–20 cm) organic carbon isotopic signatures and associated climatic factors in central-east Asia in an attempt to develop transfer functions that can be used to retrieve the paleoclimatic information stored in the thick eolian–paleosol sequences within the area. Our analysis shows that the negative correlation between the surface-soil organic δ13C values and the mean annual precipitation is robust (R2 = 0.453; n = 196; p < 0.05) and the negative correlation with the growing-season (April–September) precipitation is more significant (R2 = 0.4966; n = 196; p < 0.05). Our study further shows that the positive correlation between the surface-soil organic δ13C values and mean growing-season aridity is most significant (R2 = 0.5805; n = 196; p < 0.05). We have smoothed both the organic δ13C values and the mean growing-season aridity values using a 3-point moving-window average-filter method in an attempt to remove some of random errors and found that the positive correlation between the two is further increased (R2 =  0.7784; n =  192; p < 0.05). These robust linear relationships demonstrate their value in reconstructing paleoclimate changes in the study area. The documented climatic dependency of the surface-soil carbon isotopic composition in the study area might have resulted both from the humidity-related isotopic enrichment processes of the dominant C3 plants (stomatal conductance and photosynthetic discrimination) and from the aridity-related abundance of C4 plants (mainly Chenopodiaceae species) along the S–N bioclimatic gradient.  相似文献   

18.
Summary Using a direct Monte Carlo simulation, population growth of helper T-cells (N H) and viral cells (N v) is studied for an immune response model with an enhanced spatial inter-cellular interaction relevant to HIV as a function of viral mutation. In the absence of cellular mobility (P mob=0), the helper T-cells grow nonmonotonically before reaching saturation and the viral population grows monotonically before reaching a constant equilibrium. Cellular mobility (P mob=1) enhances the viral growth and reduces the stimulative T-cell growth. Below a mutation threshold (P c), the steady-state density of helper T-cell (p H) is larger than that of the Virus (p v); the density difference Δp o(=pV−pH) remains a constant at P mob=1 while −Δp o→0 as P mutP c at P mob=0. Above the mutation threshold, the difference Δp o in cell density, grows with ΔP=P mutP c monotonically: ΔP o ∞ (ΔP)β ≃ with β≈0.574±0.016 in absence of mobility, while Δp o≈6(ΔP) with P mob=1.  相似文献   

19.
20.
Gravimetric and a combination areal-gravimetric methods for determining dry matter budgets for leaf eating Lepidoptera were compared. The gravimetric method is based on dry weight/live weight ratios of the leaves fed to the larvae. In the areal-gravimetric method, the quantity of food offered to the larvae is determined from the area of leaf tracings and the dry matter content per unit area of the leaves. The areal-gravimetric method permits the use of larger leaf sprays and an open, gauze enclosed rearing chamber. There were no consistent differences in budget factors (growth, ingestion or egestion), nor were there any differences in the observed variability of the data attributable to the method used. However, the expected variability based on instrument precision for the gravimetric method is less than for the areal-gravimetric method. Experimental factors inherent in the gravimetric method introduce variability to the measurements that are not present in the areal method. Thirty to 60% of the variability in budget factors was attributed to intrinsic properties of the larvae, even though the larvae were taken from the same egg masses.
Résumé Les budgets en matière sèche consommée par des lépidoptères ont été comparés par les méthodes gravimétrique et planimétrique. La méthode gravimétrique est basée sur le rapport poids sec/poids frais de feuilles consommées par les chenilles. Avec la méthode planimétrique, la quantité d'aliment proposée aux chenilles est déterminée par les tracés de la surface des feuilles et le contenu de matière sèche par unité de surface des feuilles. La méthode de planimétrie permet l'utilisation de plus grands rameaux de feuilles et de cages d'élevage extérieures en gaze. Il n'y avait pas de différence appréciable dans les éléments du budget (croissance, ingestion et déjection), ni aucune différence dans la variabilité observée des données attribuable à la méthode utilisée. Cependant, la variabilité attendue d'après la précision des mesures avec la méthode gravimétrique est inférieure à celle de la méthode planimétrique. est inférieure à celle de la méthode planimétrique. Des éléments expérimentaux, inhérents à la méthode gravimétrique, introduisent une variabilité dans les mesures que l'on n'a pas avec la méthode planimétrique. 30–60% de la variabilité dans la consommation ont été attribués à des paramètres internes à la chenille, même quand elles provenaient toutes de la même ooplaque.
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