Autolyse der zellwand bei den gameten von Chlamydomonas reinhardii

Zusammenfassung An der Gametenverschmelzung bei der heterothallischen, isogamen Grünalge Chlamydomonas reinhardii ist, wie an der Befruchtung bei höheren Organismen, ein lytischer Faktor beteiligt. Während des Kontaktes von und Gameten oder während einer, Isoagglutination von Gameten des einen Paarungstyps mit den isolierten Geißeln vom entgegengesetzten Paarungstyp wurde ein hitzelabiler, lytischer Faktor ins Nährmedium abgegeben. Auch Extrakte aus vegetativen und generativen Zellen von C. reinhardii enthielten lytische Aktivität. Unter der Einwirkung des extrahierten oder des sekretierten Autolysins fand eine partielle Lyse der Zellwände von Gameten und Zoosporen statt, die mit der Freisetzung von Protoplasten verbunden war. Die Präparate mit lytischer Aktivität lösten außerdem die Wände der Zoosporangien von C. reinhardii vollständig auf und setzten dabei Zoosporen frei.

---

Autolysis of the cell wall from the gametes of Chlamydomonas reinhardii

Summary As in fertilization of higher organisms a lytic factor is involved in the mating reaction of the heterothallic isogamous green alga Chlamydomonas reinhardii. Lytic activity was found in the medium after copulation of and gametes, after isoaggllutination of gametes with isolated flagella, and of gametes with isolated flagella. A lytic factor could also be extracted from vegetative and generative cells of C. reinhardii. Partial lysis of cell walls from vegetative or generative cells accompanied by the release of protoplasts, and complete lysis of the walls from zoosporangia followed by the release of zoospores, were observed in the presence of the autolysin secreted by gametes under the above mentioned conditions or extracted from vegetative and generative cells.

     

Measurements of electrical potential differences across yeast plasma membranes with microelectrodes are consistent with values from steady-state distribution of tetraphenylphosphonium inPichia humboldtii

Summary Electrical potential differences across the plasma membrane () of the yeastPichia humboldtii were measured with microelectrodes (filled with 0.1m KCl) inserted into cells immobilized in microfunnels. The registered signals were reproducible and stable for several minutes. On attainment of stable reading for the specific membrane resistanceR _sp was determined by applying square-current pulses to the preparation. Both andR _sp were pH dependent and displayed equal but opposite deflection, reaching its maximal value of –88±9 mV (n=13) andR _sp its minimal value of 10 k·cm² (maximal conductance) at pH 6.5. Uncouplers and the polyene antibiotic nystatin depolarized the cells, decreasing to –21±15 mV (n=10) with concomitant decrease ofR _sp. Comparison of values from microelectrode measurements with those calculated from the steady-state distribution of tetraphenylphosphonium ions agreed within 10 mV under all physiological conditions tested, except at pH values above 7.0. During microelectrode insertion transient voltage signals (a few msec long) were detected by means of an oscilloscope. These voltage signals were superimposed on the stable recordings described above. These short voltage signals disappeared in uncoupled cells. The closely related values obtained by two independent methods (direct measurements with microelectrodes and calculation from steady-state distribution of a lipophilic cation) provide evidence that these values reffect the true membrane potential of intact cells.     

Observations on the morphogenetic capacity of “residual nuclei” inAcetabularia

Summary Stalks ofAcetabularia mediterranea cells after cyst formation contain residual nuclei,e.g., secondary nuclei not used for cyst formation. Residual nuclei may lead to the formation in the stalk of direct germlings (Hämmerling 1955), cysts, and—without preceeding cyst formation—of biflagellate swarmers, identical in appearance with gametes.     

The energy transmission in ATP synthase: From the γ-c rotor to the α3β3 oligomer fixed by OSCP-b stator via the βDELSEED sequence

ATP synthase (F₀F₁) is driven by an electrochemical potential of H⁺ (H⁺). F₀F₁ is composed of an ion-conducting portion (F₀) and a catalytic portion (F₁). The subunit composition of F₁ is ₃₃. The active ₃₃ oligomer, characterized by X-ray crystallography, has been obtained only from thermnophilic F₁ (TF₁). We proposed in 1984 that ATP is released from the catalytic site (C site) by a conformational change induced by the DELSEED sequence via -F₀. In fact, cross-linking of DELSEED to stopped the ATP-driven rotation of in the center of ₃₃. The torque of the rotation is estimated to be 420 pN·å from the H⁺ and H⁺-current through F₀F₁. The angular velocity () of is the rate-limiting step, because H⁺ increased theV _max of H⁺ current through F₀, but not theK _m (ATP). The rotational unit of F₀ (=ab₂c₁₀) is /5, while that in ₃₃ is 2/3. This difference is overcome by an analog-digital conversion via elasticity around DELSEED with a threshold to release ATP. The distance at the C site is about 9.6 å (2,8-diN₃-ATP), and tight Mg-ATP binding in ₃₃ was shown by ESR. The rotational relaxation of TF₁ is too rapid (=100 nsec), but the rate of AT(D)P-induced conformational change of ₃₃ measured with a synchrotron is close to . The ATP bound between the P-loop and E188 is released by the shift of DELSEED from RGL. Considering the viscosity resistance and inertia of the free rotor (-c), there may be a stator containing OSCP (= of TF₁) and F₀-d to hold free rotation of ₃₃.     

Ultrahistochemical study on the ruthenium red surface staining

Summary The cell coat picture effect which is usually obtained with the conventional RR method, that is with the RR/OsO₄ coupled reaction, is investigated. In this first paper, each of conceivable events which might take place between RR, OsO₄ and cell surface membrane is discussed or studied. Various tests are carried out on ascites Ehrlich carcinoma cells and Zajdela ascites hepatoma cells treated with numerous chemical reagents, as also on a few pure proteins. The set of data supports the concept that the staining pattern is due to the combination in surface membranes of RR with a colloidal-like form of OsO₂. The latter might occur during the formation of stable cyclic osmic acid diesters between OsO₄ and membrane unsaturated lipids. A possibility by which the resulting marker is though also to be in a colloidal-like state is put forward. A next report will deal with this problem.     

Three-dimensional organization of the plasmalemmal vesicular system in directly frozen capillaries of the rete mirabile in the swim bladder of the eel

Summary Several recent studies comparing chemically fixed and cryofixed endothelium have indicated that glutaraldehyde fixation may result in increases in the population of vesicles in the cytoplasm. Other reports based on ultrathin serial-section reconstruction of chemically fixed endothelium have revealed that the vesicular system is comprised of interconnected membranous compartments, which are ultimately continuous with either cell surface but do not extend across the endothelial cell. In this study, we have investigated the three-dimensional organization of the vesicular system in directly frozen, freeze-substituted capillaries of the rete mirabile from the swim bladder of the eel, specifically using the same block of embedded capillaries in which frozen capillaries had previously been found to contain less vesicles than chemically fixed capillaries. The results show that essentially all vesicles remain inter-connected with each other and are part of two separate sets of invaginations from the luminal and abluminal cell surface like in chemically fixed tissue. Any increase in vesicle number resulting from glutaraldehyde fixation does not affect the overall three-dimensional organization of the vesicular system in these endothelial cells.     

Lipid profile of rat myelin subfractions

Myelin from adult rat brains was separated on a discontinuous sucrose gradient into three subfractions. Analysis of light, heavy and membrane fraction lipid classes was performed by HPTLC and densitometry while fatty acid composition was determinated by GLC. The more interesting results observed are: i) the membrane fraction resembles in its lipid and fatty acid composition other cell membranes (particulary oligodentrocytes); ii) light and heavy myelin are quite similar between them but the former has a higher content of sphingomyelin, a lower hydroxy/nohhydroxy cerebrosides ratio and a lower content of monoenoic fatty acids than the heavy subfraction. The results obtained could explain the different structures observed in each myelin subfraction since fatty acid composition, hydroxy fatty acids, sphingomyelin and cholesterol play a key role in the stability and structure of membranes.     

Coral growth in high-nutrient,low-pH seawater: a case study of corals cultured at the Waikiki Aquarium,Honolulu, Hawaii

Fifty-seven species of hermatypic corals have been maintained and grown in high-nutrient seawater at the Waikiki Aquarium, Honolulu, Hawaii. In this study we document the chemical conditions of aquarium water in terms of dissolved nutrients and carbon. Aquarium water is characterized by concentrations of inorganic nutrients that are high relative to most natural reef ecosystems: SiO₃ 200 M; PO₄ 0.6 M; NO₃ 5 M; NH₄ 2 M. In contrast, concentrations of organic nutrients are lower than most tropical surface ocean waters: DOP 0.1 M and DON 4 M. The incoming well-water servicing the facility has low pH, crating over-saturation of carbon dioxide. The coral communities in aquaria took up inorganic nutrients and released organic nutrients. Rates of nutrient uptake into aquaria coral communities were similar to nutrient uptake by natural reef communities. Coral growth rates were near the upper rates reported from the field, demonstrating corals can and do flourish in relatively high-nutrient water. The growth of corals does not appear to be inhibited at concentrations of nitrogen up to 5 M. Statements implying that corals can only grow in low nutrient oligotrophic seawater are therefore oversimplifications of processes that govern growth of these organisms. Some basic guidelines are given for maintenance of coral communities in aquaria.     

Primary structure of the hemoglobin beta-chain of rose-ringed parakeet (Psittacula krameri)

The primary structure of Rose-ringed Parakeet hemoglobin -chain was established, completing the analysis of this hemoglobin. Comparisons with other avian -chains show variations smaller than those for the corresponding -chains. There are 11 amino acid exchanges in relationship to the only other characterized psittaciform -chain, and a total of 35 positions are affected by differences among all avian -chains analyzed (versus 61 for the -chains). At three positions, the Psittacula -chain has residues unique to this species. Three ₁₁ contacts are modified, by substitutions at positions 51, 116, and 125.     

Modulation of formation of tumor metastases by peritoneal macrophages elicited by various agents

Summary We have studied the formation of experimental B16 melanoma metastases in the lungs of mice inoculated IV with tumoricidal or nontumoricidal peritoneal macrophages elicited by various agents. IV inoculation of peritoneal M elicited by Brewer's thioglycollate medium (TG-M) 1 day before the injection of B16 melanoma cells dramatically increased the number of metastatic foci in the lungs. NIH thioglycollate broth and proteose peptone each elicited a relatively low number of M, which were morphologically distinguishable from TG-M and did not influence the yield of B16 melanoma colonies in the lungs. Resident or C. pravum-elicited M also did not augment metastatis formation. TG-M became highly tumoricidal after IP stimulation with poly I: C. However, tumoricidal TG-M inoculated IV 1 day before IV inoculation of B16 melanoma cells did not have an antimetastatic effect. On the contrary, both tumoricidal and nontumoricidal TG-M augmented metastasis formation. Poly I: C treatment had a substantial antimetastatic effect in the normal mice, but not in mice with adoptively transferred TG-M. Histological analysis revealed that IV-inoculated TG-M (tumoricidal or nontumoricidal, either viable or disrupted) induced severe intravascular reaction in the lungs, but not in the liver or kidney. This reaction manifested in the aggregation of the various blood cells, preferentially neutrophils. These reactions were not observed after IV inoculation of PM or NIH TG-M.Intravascular inflammatory reactions induced by TG-M may be responsible for the augmentation of metastasis formation, partly by suppression of NK reactivity and mostly by the acceleration of the processes of tumor cell extravasation. These data may provide some insight into the failure to achieve systemic adoptive immunotherapy using activated peritoneal TG-M. Abbreviations used in this paper are: TG-M, thioglycollate-elicited macrophages; PM, proteose-peptone-elicited macrophages; NIH TG-M, macrophages elicited with NIH thioglycollate broth; CP-M, macrophages; elicited with C. parvum; poly I: C, polyinosinic: polycytidylic acid; TGM, thioglycollate medium; NIHTGB, NIH thioglycollate broth     

Ganglioside lactones:1H-NMR determination of the inner ester position of GD1b-ganglioside lactone naturally occurring in human brain or produced by chemical synthesis

The complete definition of the chemical structure of G_D1b-ganglioside (G_D1b) lactone isolated from human brain has been given by means of spectrometric and spectroscopic analyses. G_D1h lactone contains a single ester linkage involving the external sialic acid carboxyl group and the C-9 hydroxyl group of the internal sialic acid unit. A synthetic lactone of G_D1b prepared treating G_D1b with glacial acetic acid characterized in the same way showed an identical chemical structure.Abbreviations: Ganglioside nomenclature is according to Svennerholm [16] and the IUPAC-IUB Recommendations [17] G_M1 G_M1-ganglioside, II³NeuAc-GgOse₄Cer, Gal1-3GalNac1-4[NeuAc2-3]Gal1-4Glc1-1Cer - G_D1b G_D1b-ganglioside, II³(NeuAc)₂GgOse₄Cer, Gal1-3GalNAc1-4[NeuAc2-8NeuAc2-3]Gal1-4Glc1-1Cer - G_D1b lactone G_D1b-L, Gal1-3GalNAc1-4[NeuAc(1-9)2-8NeuAc2-3]Gal1-4Glc1-1Cer - Cer ceramide - FAB-MS fast atom bombardment-mass spectrometry - ¹H-NMR proteon nuclear magnetic resonance - 1D-NMR one dimensional NMR - 2D-COSY two dimensional correlated spectroscopy - DMSO-d₆ deuterated dimethylsulfoxide     

Tubulin Stability and Decay: Mediation by Two Distinct Classes of IKP104-Binding Sites

IKP104, a novel antimitotic drug, has two classes of binding sites on bovine brain tubulin with different affinities. IKP104, by itself, enhances the decay of tubulin, but in the presence of colchicine or podophyllotoxin, it stabilizes tubulin instead of opening up the hydrophobic areas [Luduena et al. (1995), Biochemistry 34, 15751–15759], Here, we have dissected these two apparently contradictory effects of IKP104 by cleaving the C-terminal ends of both and subunits of tubulin with subtilisin. We have found that the selective removal of the C-terminal ends from both the and subunits of tubulin lowers the sulfhydryl titer by approximately 1.5 mol/mol of dimer. Interestingly, IKP104 does not increase either the sulfhydryl liter or the exposure of hydrophobic areas of this subtilisin-treated tubulin (_ss). Moreover, IKP104 lowers the sulfhydryl titer of _ss tubulin approximately by 1 mol/mol and appears to inhibit completely the time-dependent decay of _ss tubulin. The cleavage at the C-terminal ends of both and modulates the effect of IKP104 on the subunit, but not on the subunit. Fluorometric binding data analysis suggests that IKP104 binds to the _ss tubulin only at the high-affinity site; the low-affinity site(s) disappear almost completely. The sulfhydryl titer data for and and the fluoromelric data therefore suggest that the interaction of IKP104 at the high-affinity site on tubulin is not regulated by the C-terminal domains of and and the effect of the high-affinity site is restricted largely to the subunit, while the low-affinity-site binding is modulated by the C-terminal domain of . It also appears that the stabilization and the acceleration of the decay of tubulin are mediated by distinct interactions of IKP104 with its high- and low-affinity sites on tubulin, respectively.     

Evidence for disaggregation of oligomeric G(o)alpha induced by guanosine-5;-3-O-(thio)triphosphate activation

Myristoylated G_o was expressed in and highly purified from Escherichia coli strain JM109 cotransformed with pQE60 (G_o) and pBB131 (N-myristoyltransferase, NMT). Non-denaturing gel electrophoresis and gel filtration analysis revealed that the G_o, in its GDP-bound form, could form oligomers involving dimer, trimer, tetramer, pentamer, or hexamer and guanosine 5"-3-O-(thio)triphosphate (GTPS) activation induced disaggregation of the G_o oligomers to monomers. The G_o was crosslinked by a cross-linker, N,N"-1,4-phenylenedimaleimide (p-PDM), yielding multiple crosslinked products. In contrast, no obvious cross-linking occurred when G_o was pretreated with GTPS. Immunoblot analysis also demonstrated oligomerization of the purified G_o proteins and its disaggregation triggered by GTPS. These results provided direct evidence for the disaggregation–coupling theory and the disaggregation action of GTPS may further elucidate the regulatory role of GDP/GTP exchange in G protein-coupled signal transduction pathways.     

Synergistic effect of BMP-2 and ascorbate on the phenotypic expression of osteoblastic MC3T3-E1 cells

Of TGF- superfamily proteins, BMP-2 enhanced alkaline phosphatase (ALP) activity in cultured osteoblastic cells, MC3T3-E1, to the same level promoted by ascorbate, whereas TGF-s (1, 2, 3) reduced ALP activity and altered cell morphology under the same conditions. Activin appeared to have no distinct effect on ALP synthesis. Ascorbate dependent increase in ALP synthesis was markedly stimulated in the presence of BMP-2. The synergistic effect of ascorbate on ALP synthesis was replaced by type I collagen coated on the culture dish. However, BMP-2 appeared not to bind to type I collagen. These findings indicate that BMP-2 acts directly on osteoblastic cells through its receptors and collagenous matrix can neither recruit BMP-2 nor modulate directly the action of BMP-2 in the pericellular area. Treatment of cells grown in ascorbate media with TGF-s decreased rapidly the cellular ALP activity indicating that TGF-s direct cells to the differentiated stage.     

A general strategy for the isolation of carbohydrate chains fromN-,O-glycoproteins and its application to human chorionic gonadotrophin

For the structural analysis of the carbohydrate chains ofN-,O-glycoproteins a straightforward strategy was developed based on the cleavage of theN-linked chains with immobilized peptide-N ⁴-(N-acetyl--glucosaminyl) asparagine amidase-F (PN-Gase-F) fromFlavobacterium meningosepticum, followed by alkaline borohydride treatment of the remainingO-glycoprotein material. This methodology was applied to the isolation of the Asn- and Ser-linked carbohydrate chains of human chorionic gonadotrophin. The structures of the isolated oligosaccharides were verified by 500-MHz¹H-NMR spectroscopy. The Asn-linked sugar chains were shown to be: NeuAc2-3Gal1-4GlcNAc1-2Man1-6[NeuAc2-3Gal1-4GlcNAc1-2Man1-3]Man 1-4GlcNAc1-4[Fuc1-6]_0-1GlcNAc and Man1-6[NeuAc2-3Gal1-4GlcNAc1-2Man 1-3]Man1-4GlcNAc1-4GlcNAc. Also some minor constituents occurred. The structures of the Ser-linked oligosaccharides were established in the form of their oligosaccharide-alditols as: NeuAc2-3Gal1-3[NeuAc2-6]GalNAc, NeuAc2-3Gal 1-3GalNAc and NeuAc2-3Gal1-3[NeuAc2-3Gal1-4GlcNAc1-6]GalNAc.Abbreviations hCG human chorionic gonadotrophin - hCG- -subunit - hCG- -subunit - ElA enzyme immunoassay - PNGase-F peptide-N ⁴-(N-acetyl--glucosaminyl)asparagine amidase-F (EC 3.5.1.52) - SDS sodium dodecyl sulphate - GalNAc N-acetylgalactosamine - GlcNAc N-acetylglucosamine - NeuAc N-acetylneuraminic acid - Man mannose - Gal galactose - Fuc fucose     

Nuclear Overhauser effect investigation on GM1 ganglioside containingN-glycolyl-neuraminic acid (II3Neu5GcGgOse4Cer)

The conformational properties of the oligosaccharide chain of GM1 ganglioside containingN-glycolyl-neuraminic acid, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer, were studied through NMR nuclear Overhauser effect investigations on the monomeric ganglioside in dimethylsulfoxide, and on mixed micelles of ganglioside and dodecylphosphocholine in water. Several interresidual contacts for the trisaccharide core--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-were found to fix the relative orientitation of the three saccharides, while the glycosidic linkage of the terminal -Gal-was found to be quite mobile as the -Gal-(1-3)--GalNAc-disaccharide exists in different conformations. These results are similar to those found for two GM1 gangliosides containingN-acetyl-neuraminic acid and neuraminic acid [1].Abbreviations Ganglioside nomenclature is in accordance with Svennerholm [23] and the IUPAC-IUB Recommendations [24] GM3(Neu5Ac) II³Neu5AcLacCer, -Neu5Ac-(2-3)--Gal-(1-4)--Glc-(1-1)-Cer - GM3(Neu5Gc) II³Neu5GcLacCer, -Neu5Gc-(2-3)--Gal-(1-4)--Glc-(1-1)-Cer - GM1(Neu5Ac) II³Neu5AcGgOse₄Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - GM1(Neu5Gc) II³Neu5GcGgOse₄Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu5Gc-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - GM1(Neu) II³NeuGgOse₄Cer, -Gal-(1-3)--GalNAc-(1-4)-[-Neu-(2-3)]--Glc-(1-1)-Cer - GD1a IV³Neu5AcII³Neu5AcGgOse₄Cer, -Neu5Ac-(2-3)--Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - GalNAc-GD1a IV⁴GalNAcIV³Neu5AcII³Neu5AcGgOse₄Cer, -GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-3)--GalNAc-(1-4)-[-Neu5Ac-(2-3)]--Gal-(1-4)--Glc-(1-1)-Cer - Neu neuraminic acid - Neu5Ac N-acetyl-neuraminic acid - Neu5Gc N-glycolyl-neuraminic acid - Cer ceramide     

The αβ complexes of ATP synthase: the α3β3 oligomer and α1β1 protomer

The basic structures of the catalytic portion (F₁, ₃₃) of ATP synthase are the ₃₃ hexamer (oligomer with cooperativity) and ₁₁ heterodimer (protomer). These were reconstituted from the and subunits of thermophilic F₁ (TF₁), and the ₃₃ hexamer was crystallized. On electrophoresis, both the dimer and hexamer showed bands with ATPase activity. Using the dimer and hexamer, we studied the nucleotide-dependent rapid molecular dynamics. The formation of the hexamer required neither nucleotide nor Mg. The hexamer was dissociated into the dimer in the presence of MgADP, while the dimer was associated into the hexamer in the presence of MgATP. The hexamer, like mitochondrial F₁ and TF₁, showed two kinds of ATPase activity: one was cooperative and was inhibited by only one BzADP per hexamer, and the other was inhibited by three BzADP per hexamer.     

Functional sites in F1-ATPases: Location and interactions

This review focuses on the location and interaction of three functional sites in F₁-ATPases. These are catalytic sites which are located in subunits, noncatalytic nucleotide-binding sites which are located at interfaces of and subunits and modulate the hydrolytic activity of the enzyme, and a site that binds inhibitory amphipathic cations which is at an interface of and subunits. The latter site may participate in transmission of conformational signals between catalytic sites in F₁ and the proton-conducting apparatus of F₀ in the intact ATP synthases.     

Charles Darwin's theory of evolution: A review of our present understanding

The paper characterizes Darwin's theory, providing a synthesis of recent historical investigations in this area. Darwin's reading of Malthus led him to appreciate the importance of population pressures, and subsequently of natural selection, with the help of the wedge metaphor. But, in itself, natural selection did not furnish an adequate account of the origin of species, for which a principle of divergence was needed. Initially, Darwin attributed this to geographical isolation, but later, following his work on barnacles which underscored the significance of variation, and arising from his work on botanical arithmetic, he supposed that diversity allowed more places to be occupied in a given region. So isolation was not regarded as essential. Large regions with intense competition, and with ample variation spread by blending, would facilitate speciation. The notion of place was different from niche, and it is questioned whether Darwin's views on ecology were as modern as is commonly supposed. Two notions of struggle are found in Darwin's theory; and three notions of variation. Criticisms of his theory led him to emphasize the importance of variation over a range of forms. Hence the theory was populational rather than typological. The theory required a Lamarckian notion of inheritable changes initiated by the environment as a source of variation. Also, Darwin deployed a use/habit theory; and the notion of sexual selection. Selection normally acted at the level of the individual, though kin selection was possible. Group selection was hinted at for man. Darwin's thinking (and also the exposition of his theory) was generally guided by the domestic-organism analogy, which satisfied his methodological requirement of a vera causa principle.     

Conformational transitions in closed circular DNA molecules I. Topological and energetical considerations

A theory of conformational transitions in closed circular DNA as a function of topological linking number of the molecule () is elaborated taking into account topological and energetical considerations. The theory predicts a step-like dependence of a number of superhelical turns in DNA molecules () on . Thus, the number of superhelical turns = for small values of . For a large (when conformational transitions begin to occur) =–_ij, where _ij is the total angle of conformational transitions for a given . This prediction is in good agreement with published data on the dependence of the sedimentation coefficient of circular DNA molecules on their topological linking number. The results also allow to explain the disagreement between a number of titratable superhelical turns in circular DNA molecules and a number of supercoiles seen on electron micrographs for molecules with sufficiently large .