Single Macromolecule Research Articles

Notes on the theory of adsorption of a single macromolecule

Three problems in the theory of adsorption of a polymeric chain are discussed in the present paper. It is shown that the phase transition during the adsorption of a macromolecule without volume effects should be treated as the passage through a tricrital point along the lines of the phase transition. Adsorption of a chain on to the surface of a cylinder of radius R is discussed. It is shown that, as R increases, the temperature of adsorption always reaches a value characteristic of adsorption on to a plane before the step increase in heat capacity occurs. The limits of applicability of the ideal chain model for adsorption on to a plane, line and point have been analysed. It is shown that, in the case of adsorption on to a surface or line, volumetric effects exist under the same conditions as in a free coil.

Computer Simulations of Complex Chemical Systems: Solvation of DNA and Solvent Effects in Conformational Transitions

As is known, atomic and small molecular systems can be realistically simulated with quantum-mechanical models. In complex chemical systems, however, the natural parameters for a description are not only the electronic density and energy but also entropy, temperature, and time. We have considered as an example for a complex chemical system the structure of water surrounding DNA with counterions. No direct experimental determination of the solvent structure around a single DNA macromolecule is available from experimental data, despite continuous efforts during the last twenty years to obtain both single crystals of DNA and scattering data from solutions. A very detailed representation is now available concerning the structure and interaction energy of water molecules in the first solvation shell or in the “grooves” of the DNA. The data obtained by our computer simulation are in good agreement with indirect data from DNA fibers at different relative humidities and with other indirect evidence. In addition, our simulated results allow us to present a preliminary model for the solvent effects in transition processes between different DNA conformations. The model presented is also in agreement with available experimental data. Finally, our results report the first determination of the position of counterions in DNA at different relative humidities and at room temperature. This application demonstrates the flexibility of the computational approach. The method we have used is very general; namely, it is not limited to a biological system but is valid for any organic or inorganic problem requiring systematic simulation on a class of compounds interacting either as a few molecules or as a large ensemble of molecules.

Isolation and characterization of the proteoglycans and collagens synthesized by cells in culture

Methods are described for isolating and characterizing the proteoglycans and collagens synthesized by cells or tissues in culture. Previous studies have established that there are different proteoglycans and collagens in the body. Disorders may involve changes in a single specific macromolecule or several.

Swelling and collapse of polymer networks

The swelling of a polymer network in a good solvent and of the collapse of this sample in a poor solvent is discussed theoretically for networks prepared in the presence of a large amount of diluent. The notation used is convenient for comparison with the theory of the swelling and collapse of a single macromolecule. The reference state for the networks under consideration must be chosen near the θ temperature for the corresponding linear macromolecule. For a free network and a network with fixed dimensions along one or two axes we obtain asymptotic results for the swelling of the network in a good solvent; these do not coincide with the classical Flory results. Results for the collapse of the network, which takes place upon cooling below the θ-point are also obtained. The collapse can occur either as a discrete first order phase transition (if the network chains are stiff or if it is stretched along one or two axes), or as a continuous non-phase transition (located in a narrow temperature range).

Observation of unstained biological macromolecules with the STEM

We discuss the observation of an unstained single biological macromolecule of human serum low density lipoprotein in a scanning transmission electron microscope (STEM). Using a carbon substrate with 16 Å average thickness and an electron dose of 19 electrons/Å 2 the image obtained from the unstained molecule shows a similar surface substructure as that seen in a negatively stained conventional transmission electron microscope (CTEM) micrograph.

Detergents Linked to Polysaccharides: Preparation and Effects on Membranes and Cells

Residues of Triton X‐100 [C8H17‐C6H4‐(O‐CH2‐CH2)9‐10‐O‐] were bound by ether bonds to inulin, dextran, amylose, and cellulose‐yielding compounds containing 10–30, 5, 19 and 6% (w/w) of Triton X‐100 residue respectively. The water‐soluble inulin derivative was studied in detail. This compound was fractionated on the basis of molecular weight by gel chromatography and on the basis of degree of substitution by adsorption to polystyrene resin. Even the residues of Triton X‐100 which were bound to a single inulin macromolecule were able to form a micelle; in addition to these monomolecular micelles the inulin derivative was able to form polymolecular micelles as well. The inulin derivative was effective in solubilizing proteins and phospholipids from membranes of human erythrocytes and liberated reverse transcriptase activity from the membrane‐enveloped virions of murine leukemia. The Triton X‐100 inulin derivative abolished binding of the 3H‐labelled antagonist dihydroalprenolol to solubilized preparations of β‐adrenergic receptors from frog erythrocytes in a dose‐related manner similar to the inactivation produced by Triton X‐100, while digitonin, a detergent containing a bulkier hydrophobic group, did not cause inactivation. On the basis of its Triton X‐100 content, the inulin derivative was found to be less detrimental to the growth of murine erythroleukemic cells in vitro than Triton X‐100 alone when short (2–4 h) exposures were used, but this difference disappeared at longer (1–3 day) exposures. Results thus suggest that the increase in size of the hydrophilic part of the detergent brings about moderation in those effects of the detergent which are dependent on the rate of diffusion, while the solubilizing and inactivating effects of the detergent were not changed. It is probably the size of the hydrophobic part which is important in protein‐inactivating properties of the detergent.

A model for the adsorption of poly(γ‐benzyl‐L‐glutamate) from solution onto alkali‐halide substrates

AbstractThe isothermal adsorption of high‐and low‐molecular‐weight poly(γ‐benzyl‐L‐glutamate) (PBLG) onto (100) alkali‐halide surfaces has been studied with the intent of developing a model for the adsorption process. Data from both the solution and adsorbed phases were correlated in this study. The α‐helical conformation was maintained in solution and on the substrate for all molecular weights investigated. Fourier‐transform infrared spectroscopy proved to be a sensitive technique for conformational analysis of adsorbed PBLG films as thin as 200 Å. High‐molecular‐weight PBLG was adsorbed as an interwoven fibrous mat, homogeneously covering the entire substrate surface. This morphology was independent of the substrate. Adsorption of low‐molecular‐weight PBLG took on a lamellar morphology, often exhibiting epitaxial orientation in the substrate 〈100〉 directions. Quasielastic laser light scattering studies showed that low‐molecular‐weight PBLG had a much higher degree of aggregation in nonpolar solvents than did high‐molecular‐weight PBLG. The proposed adsorption model is essentially a modification of one suggested by Silberberg. It involves segmental adsorption of aggregates, as opposed to single macromolecules of PBLG. The unadsorbed segments of the aggregates form a concentrated interfacial region directly above the substrate. Further interaction and association of PBLG may occur in this region, thus reducing the epitaxial ordering influence of the substrate. Consequently, polymers which do not interact strongly in the concentrated interfacial region, such as polyethylene, are expected to be adsorbed epitaxially, as has been reported.

Multiple bands produced by interaction of a single macromolecule with carrier ampholytes during isoelectric focusing

Equilibrium isoelectric focusing patterns have been computed for reversible, carrier ampholyte-induced macromolecular isomerization reactions. The calculations predict that an amphoteric macromolecule, interacting with n species of ampholyte located at different positions along the isoelectric focusing column, can give a pattern showing n + 1 well-resolved peaks under appropriate conditions.

Particle morphology in the early stages of radiation‐induced heterophase polymerization of vinyl chloride

AbstractInvestigations of the particle morphology of poly(vinyl chloride) produced under quiescent conditions during radiation‐induced bulk polymerization over the temperature range −30 to 70°C were carried out. The observations were mainly confined to the early stages of polymerization. For polymerization temperatures below about 20°C, the systems remain predominantly homogeneous during the entire polymerization and the polymer particles increase in size linearly with conversion. At higher temperatures the polymer particles rapidly settle and become cemented together. The findings are discussed in the light of the kinetic data on vinyl chloride polymerization, and a process of particle formation and growth, resembling that recently proposed by Fitch for emulsion systems, was formulated. Primary particles are initially formed by the coiling up of single macromolecules or single macroradicals and, subsequently, they increase in size by sweeping up growing free radicals from the liquid monomer phase. The free radicals which escape capture give rise to new primary particles, but their number progressively decreases as the number of the dispersed particles increases. Simultaneously, the polymer particles undergo flocculation which in a short time results in the formation of large agglomerates. As the volume of the resulting agglomerates increases, the flocculation rate decreases and, eventually, becomes so low that the flocculation does not proceed further. At low temperatures the flocculation almost ceases when the agglomerates are still small enough for sedimentation to occur only very slowly. However, this is not the case at higher temperatures. The addition of substances such as alcohols, brings about a reduction in the flocculation rate and, hence, in the size of the agglomerates formed at the end of the flocculation process. In this way, one can also obtain at high temperatures agglomerates of small sizes which remain dispersed for a long time.

The entropy of a single Gaussian macromolecule in a noninteracting solvent

The entropy of a single Gaussian macromolecule in a noninteracting solvent

Resolution of beta-adrenergic receptor binding and adenylate cyclase activity by gel exclusion chromatography.

The frog erythrocyte membrane provides an excellent model system for the study of beta-adrenergic receptor-adenylate cyclase interactions since it possesses an adenylate cyclase enzyme which is very responsive to catecholamines. The purpose of these studies was to evaluate directly whether the functions of receptor binding and adenylate cyclase activity are carried out by a single macromolecule or separable molecular entities. Obtaining this information is a first step in understanding at a molecular level how receptor binding is "coupled" to enzyme activation. Binding and cyclase activities were solubilized from the frog erythrocyte membrane with digitonin and were observed to partition independently during gel exclusion chromatography in the presence of solubilizing detergent. This finding documents that the beta-adrenergic receptor and adenylate cyclase enzyme are, in fact, separable macromolecules. Under the particular experimental conditions employed, the elution of beta-adrenergic receptor binding on Sepharose 6B was not altered by the absence or presence of beta-adrenergic agonist or antagonist ligands or by exposure of the membranes prior to solubulization to the guanyl nucleotide analog, guanyl-5'-yl imidodiphosphate.

Intramolecular conformational transitions "random coil-helix-folded structure" in polypeptides.

AbstractA general theory has been developed for conformational intramolecular transitions in a single macromolecule with a high degree of polymerization (an infinite length model) capable of forming two types of ordered structures: the α‐helix and the folded β‐structure, as well as acquiring the random coil conformation. The phase diagram analysis of this system has shown that the regular β‐structure state is separated from all other states of the chain by the phase boundary line. Any intersection of the phase boundary is a phase transition which can be either of the first order or second order, depending on values of the energy parameters of the system. Mechanisms of intramolecular rearrangements: β‐structure–random coil and α‐helix–β‐structure have been discussed. It has been shown that there exist two different mechanisms for each of these rearrangements, and the regions of parameter variation corresponding to each mechanism have been specified.

Chemical kinetics and fluorescence correlation spectroscopy.

The dynamics of macromolecules, the subject of this symposium, are most directly studied by simply looking through a microscope and observing the molecular motion. With a microscope, we can resolve the size and shape of large particles, as well as monitor dynamic motion. For smaller particles, particularly single macromolecules, we cannot resolve the size or shape; but it is still possible to observe the motion, if we can make the particles appear as bright points of light sprinkled dilutely over a dark background. Siedentopf & Zsigmondy (1903) demonstrated this fact with a device which came to be called the ultramicroscope.

Colchicine-binding protein in rat brain: Biochemical consistency and immunological discrepancy

The colchicine-binding protein in the soluble fraction of rat brain was characterized biochemically and immunologically. On the basis of molecular size and charge, the tight binding of colchicine appeared to be the property of a single macromolecule. Colchicine-binding protein was purified from the soluble fraction of rat brain by the ammonium sulfate fractionation-DEAE chromatography method and also by precipitation with vinblastine sulfate. Both preparations interacted similarly with antibody but only the latter elicited a humoral immune response in rabbits. The possible significance and causes of the immunological discrepancy are discussed.

Composite protostructures: An exercise in model building

AbstractOne of the most evident characteristics of life, at the subcellular, macroscopic level, is the phenomenon of structure, i.e., the anisotropic organization of matter and processes in space and in time. However, if we know a good deal about the (macroscopically isotropic aspects of the) chemistry of life, our understanding of structure (as defined above) does not go very far beyond what is known about the configuration and conformation of single (macro)molecules, or of finite or infinite symmetrical assemblies of (macro)molecules, and about changes in configuration, conformation, or spatial arrangement of these units – changes to which stereospecific intramolecular and intermolecular interactions can be assigned as causes. As to other macroscopic structures (on a scale comprised between the dimensions of single macromolecules and those of cells), the belief seems to prevail that the principles governing these are of the same kind as the principles governing the former: that “stereospecific intrinsic molecular determinisms” unambiguously determine every kind of biological structure.In this article, attention is drawn to other macroscopic mechanisms for the organization of matter and processes, mechanisms which are known in classical polymer chemistry under the topics of microphase separation and phase inversion, which are phenomena proper to the macroscopic level of size, not necessarily predictable from, or reducible to, stereospecific molecular determinisms. An attempt is made, at the level of hypothesis and speculative model building, to investigate the possible significance of these phenomena for biological organization.

General dynamic theory of macromolecular networks. II. Dynamics of network deformation

AbstractOn the basis of the general concepts of a macromolecular network as defined in the preceding paper, a detailed analysis of forces acting on a network chain is given. In all types of networks the chain tension involves elastic, diffusional, contact friction and internal viscosity terms; in entanglement networks there appears also an additional term associated with chain–chain friction at entanglement junctions. The force balance equation for a tetrafunctional network junction is derived. It is shown that exact formulation of this equation requires knowledge of the simultaneous configurations of all the network chains in the system. For energetic networks, solution of the force balance equation is not required for the determination of the distribution function; and the dynamics of such networks can be discussed in terms of a distribution function for a single chain. On the other hand, for entanglement and contact networks, where the force balance equation is a source of information about sliding rates, some simplified equations can be formulated and solved, by using a 4(N + 1)‐dimensional distribution function for a single macromolecule with N network junctions. It is shown that for both network classes with nonlocalized junctions the position of a network chain within the macromolecule plays an essential role and is a source of the effects of molecular weight on the physical behavior of the system.The analysis of forces acting on a network chain reveals also some essential differences in the dynamic behavior of energetic, entanglement and contact networks and thus confirms the significance of the classification proposed in the preceding paper.

Studies of adsorption onto synthetic latices

AbstractDespite the importance of reactions at the polymer‐water interface of synthetic latices very little work has been reported on the adsorption of surfactants or water‐soluble macromolecules at this site. In the studies reported here the adsorption isotherms of sodium lauryl sulfate onto polystyrene and poly(n‐butyl methacrylate) and of hydroxyethyl cellulose onto poly(n‐butyl methacrylate) are shown to be of the Langmuir type. Other water‐soluble macromolecules and latex substrates were studied at a single macromolecule content and a single latex solids level. The effective surface area of the sodium lauryl sulfate molecule, at the critical micelle concentration, was found to be 58.5 Å2 for both polystyrene and poly(n‐butyl methacrylate). This is in good agreement with results obtained by soap titration. The adsorption of macromolecules is strongly influenced by macromolecule type as well as by the latex substrate, i.e., the type of surfactant on the latex surface as well as the polymeric composition of the latex.

Two or more electrophoretic zones from a single macromolecule.

Two or more electrophoretic zones from a single macromolecule.

Localization of intracellular polysaccharide granules in Streptococcus mitis

The accumulation of an intracellular glycogen-like polysaccharide was studied by electron microscopy in a strain of Streptococcus mitis which can store up to 50 per cent of its dry weight as intracellular polysaccharide. Storage of polysaccharide was reflected by an increase in the number but not the size of distinct cytoplasmic granules. Synthesis of glycogen appeared to occur near the cell membrane since the granules in the earlier stages of accumulation were usually located there. The average diameter of the granules was about 600 Å and from this measurement the molecular weight of the polysaccharide was estimated to be about 3.3 × 10 7. Each granule would appear to represent a single glycogen macromolecule.

THIN-LAYER SEPHADEX CHROMATOGRAPHY AND IMMUNODIFFUSION

With a few simple modifications of thin- layer chromatography on Sephadex G-200 'Superfine' gels, molecular size, amount, enzymic and/or immunological activity can be determined on 1 μl. samples containing 1 to 10 μg. single or mixed macromolecules. Extension to the submicrogramme scale is possible with radioactively labelled material and autoradiography. A convenient sampling system allows the characterisation of antigens and antibodies by a variety of tests (gel diffusion, immunoelectrophoresis, hæmagglutination or its inhibition, complement fixation, &c.).