Protein-like Copolymer Research Articles

Effect of Protein-like Copolymers Composition on the Phase Separation Dynamics of a Polymer Blend: A Monte Carlo Simulation

We use kinetic Monte Carlo simulation based on the bond fluctuation model to investigate the dynamics of phase separation in immiscible 80/20 A/B binary polymer blends, comprising 80% and 20% of A and B components, respectively, in the presence of ≈4.92% 30-mer protein-like copolymer (PLC) made of C and D segments. The molecular interactions are chosen such that there is an attraction between A and C and between B and D segments and no interaction between like segments; all other interaction energies have been chosen to be repulsive. The PLC migration to and presence at the A/B interface effectively slow down the process of phase separation in binary blends, thereby minimizing the unfavorable A/B contacts and reducing the A/B interfacial tension. The ability of PLCs to effectively retard the process of phase separation depends sensitively on the PLC composition. PLCs with 0.3 ≤ xC ≤ 0.5, where xC is the mole fraction of C, are most effective in compatibilizing the 80/20 A/B binary blend. The growth of pha...

Isolation and characterization of a novel cyanophycin synthetase from a deep-sea sediment metagenomic library

Cyanophycin is non-ribosomally synthesized protein-like copolymer. Synthesis of cyanophycin is catalyzed by cyanophycin synthetase (CphA). In this study, a novel cyanophycin synthetase CphA49 belonging to NOR5 clade of Gammaproteobacteria was identified with primer-based screening from a deep-sea sediment metagenomic library. The cphA49 gene contained an open reading frame of 2,637bp and encoded a protein with a predicted molecular mass of 100kDa. A recombinant CphA49 was obtained by the functional expression of cphA49 in Escherichia coli BL21 (DE3). The biochemical properties of the purified CphA49 were determined. The optimum pH and temperature of the recombinant CphA49 were 9.0 and 40°C, respectively. The enzyme was stable at temperatures below 40°C. The recombinant CphA49 exhibited strict primer dependency and broad substrate specificities. Cyanophycin catalyzed by CphA49 exhibited homogenous molecular mass. The amino acid composition of cyanophycin was determined and constitutes arginine, aspartic acid, and lysine.

Phase transition of a single protein-like copolymer chain

Protein-like copolymers (PLCs) are a kind of artificial macromolecule owning protein characteristics. We investigate the general phase transition of a single PLC chain with uniform block structure using a parallel Wang–Landau sampling method. Two typical PLC models, i.e. HP and AB models, are employed to reveal the hydrophobic effect and the phase separation effect for a PLC chain folding in aqueous environments. It is found that the block length m greatly influences the phase transition of PLC. With increasing m, the low energy stable structures for the HP model change from a tube-like aggregation to a tadpole-shape structure, and for the AB model they evolve from a multilayer structure to two separated spheres. When m is very small, the liquid–crystal transition disappears. When m is larger, the AB-PLC shows two first order transitions corresponding to the A and B phase separation transition and the liquid–crystal transition, respectively, while the HP model only shows one first order liquid–crystal transition. We further found that during the freezing of the AB-PLC, the whole chain experiences a special intermediate state where one component is embedded by another component. The result is valuable to design the functional single-molecule devices and next generation nano building blocks using PLCs.

Phase Separation Dynamics for a Polymer Blend Compatibilized by Protein-like Copolymers: A Monte Carlo Simulation

We use Monte Carlo simulation based on the bond fluctuation model to investigate how adding ≈4.92% protein-like copolymer (PLC) to an immiscible binary polymer blend affects the dynamics of phase separation. PLCs slow down effectively the process of phase separation in binary blends by migrating to the biphasic interface between the immiscible homopolymers, thereby reducing the interfacial tension. The ability of PLCs to retard effectively the process of phase separation depends sensitively on the interaction energy between the PLCs and homopolymers and the PLC chain length. PLCs compatibilize the binary blend more effectively with increasing attractive interaction between the PLC blocks and homopolymers. Nominal improvement in compatibilization of the binary blend is achieved with increasing PLC chain length. The growth of phase-separated domains follows a dynamical scaling law for both the binary blend and PLC compatibilized ternary blend in the late stages of phase separation. The universal scaling functions are nearly independent of the interaction energy and PLC chain length. Thus, the phase-separated domains grow with dynamical self-similarity irrespective of the type of PLC added to the binary blend, although the type of PLC significantly alters the growth rate of the phase-separated domains.

Energetics of the binding of Cu(II) ions by thermosensitive copolymers of N-vinylcaprolactam and N-vinylimidazole in different conformational states of macromolecules

Random and protein-like copolymers based on N-vinylcaprolactam and N-vinylimidazole are synthesized by free radical polymerization in an aqueous solution. The above copolymers show a different thermal behavior in aqueous media at pH 7.2. At 45°C, the solution of a random copolymer experiences phase separation, whereas a protein-like copolymer undergoes a transition from the unfolded conformation to the compact conformation without any phase separation. The method of isothermal titration calorimetry is used to study the binding of Cu(II) ions by protein-like and random copolymers of N-vinylcaprolactam and N-vinylimidazole at 25 and 45°C, which correspond to different conformational states of macromolecules. The standard enthalpy and constant of binding are estimated. For both copolymers, the enthalpies of binding are negative and similar. When temperature is increased from 25 to 45°C, the constant of binding of copper ions by a protein-like copolymer increases by more than three orders of magnitude, whereas the corresponding constant of a random copolymer remains almost unchanged. Therefore, the transition of protein-like copolymer from the coiled conformation to the compact conformation noticeably facilitates the formation of an imidazole quasi-receptor, which is characterized by a certain spatial configuration and by a high affinity for the functional ligand. This effect is provided by an entropy gain no less than 50 J/(mol K).

Destruction of globules of Co- and homopolymer macromolecules in the presence of an amphiphilic substrate

The destruction of a globule in the presence of a dimeric substrate composed of a hydrophilic group OP and a hydrophobic group H with a high affinity to hydrophobic H units of a macromolecule has been studied. Globules of the homopolymer H macromolecule and the macromolecule of the HP copolymer with proteinlike statistics of monomer unit distribution along a chain have been investigated. The destruction of a globule in such systems begins with the transformation of the globule’s shape from spherical to disklike. At high substrate concentrations, the globule of the proteinlike copolymer is completely destroyed; under the same conditions, the homopolymer macromolecule forms a structure composed of two beads having a shape close to that of the oblate ellipsoid that are located symmetrically about a string connecting them.

Catalytic properties of the protein-like copolymer of N-vinylcaprolactam and N-vinylimidazole in the hydrolysis of an ester substrate

Catalytic properties of the protein-like copolymer of N-vinylcaprolactam and N-vinylimidazole in the hydrolysis of an ester substrate

Core/Shell Structures of Proteinlike Copolymers: Are Finite Aggregates Thermodynamically Stable?

We propose a theory of aggregation in solutions of amphiphilic copolymers consisting mostly of insoluble H (hydrophobic) units with a small fraction of soluble (P, polar) monomer units. When P units are arranged along the sequence in a periodic (regular) fashion, the resultant HP copolymers are essentially insoluble: they precipitate. The main result is that finite aggregates of HP copolymers can be made stable by an appropriate smart arrangement of the same number of soluble P units in the chemical sequence. An analytical approach yielding the thermodynamic quantities of H-core/P-shell copolymer structures in the weak stretch limit is developed. We show that different types of copolymer aggregates and microdomain structures can be thermodynamically stable depending on the copolymer chemical sequence. The relationship between the block-length distribution and copolymer aggregation is illustrated by a few phase diagrams. Copolymers that can form stable finite aggregates with H-core/P-shell structure (including single-chain globules) are often referred to as the proteinlike copolymers. Thus, the present theory sheds a new light as to the essential features of proteinlike copolymer sequences.

Computer modeling of synthesis of proteinlike copolymer via copolymerization with simultaneous globule formation

Using a Monte Carlo simulation technique, we have modeled the process of copolymerization of hydrophobic and hydrophilic monomers in a selective (polar) solvent. The composition of an emerging polymer chain is such that macromolecule adopts a globular conformation. The preferential sorption of hydrophobic monomers in the core of the globule is explicitly taken into account. It is shown that such the copolymerization process automatically leads to the formation of the core–shell microstructure in the resulting globule and to the well-pronounced long-range correlations of the Levy-flight type in obtained sequences of monomer units. Thus, this type of synthesis provides a robust one-step method of producing of “proteinlike” copolymers, i.e., copolymers that exhibit in the globular state a microstructure with a hydrophobic core wrapped in a hydrophilic envelope.

Binding of Cu(II)-Poly(N-isopropylacrylamide/vinylimidazole) Copolymer to Histidine-Tagged Protein: A Surface Plasmon Resonance Study

The thermoresponsive copolymer of N-isopropylacrylamide (NIPAM) with 1-vinylimidazole (VI), poly(NIPAM-VI), synthesized by radical polymerization has been used to purify the histidine-tagged green flourescent protein (His-tag GFP) from recombinant E. coli by metal-chelate affinity precipitation. The purified protein was immobilized on the BIAcore sensor chip by carbodiimide coupling. Affinity binding of the Cu(II)-loaded copolymer, poly(NIPAM-VI), to the His-tag GFP-immobilized surface was monitored by surface-plasmon-resonance (SPR) measurements. Complete recovery of the metal copolymer from the surface was achieved with either using the monomer displacer, 200 mM imidazole buffer, or the polymeric displacer, copolymer of poly(NIPAM-VI) (26 mol % VI). The conformation of the copolymer was a critical factor for the metal interactions and hence displacement of the metal copolymer. With the proposed conformation of protein-like copolymers (Wahlund, P.-O.; Galaev, I. Yu.; Kazakov, S. A.; Lozinsky, V. I.; Mattiasson, B. Macromol. Biosci. 2002, 2, 33.), the SPR study confirmed the prediction of exposed imidazole groups in the poly(NIPAM-VI). The complete elution of the affinity-bound metal copolymer was achieved with protein-like copolymer (imidazole groups exposed to the outer solution), and no recovery was obtained with IMAC nonbound copolymer fraction (imidazole groups unexposed). The SPR measurement showed a sharp phase transition of affinity adsorbed thermoresponsive Cu(II)-poly(NIPAM-VI) copolymer at 32 °C, thus proposing a sensitive way to determine lower critical solution temperature.

Coil-globule transition for regular, random, and specially designed copolymers: Monte Carlo simulation and self-consistent field theory

The coil-globule transition has been studied for A-B copolymer chains both by means of lattice Monte Carlo (MC) simulations using bond fluctuation algorithm and by a numerical self-consistent-field (SCF) method. Copolymer chains of fixed length with A and B monomeric units with regular, random, and specially designed (proteinlike) primary sequences have been investigated. The dependence of the transition temperature on the AB sequence has been analyzed. A proteinlike copolymer is more stable than a copolymer with statistically random sequence. The transition is more sharp for random copolymers. It is found that there exists a temperature below which the chain appears to be in the lowest energy state (ground state). Both for random and proteinlike sequences and for regular copolymers with a relatively long repeating block, a molten globule regime is found between the ground state temperature and the transition temperature. For regular block copolymers the transition temperature increases with block size. Qualitatively, the results from both methods are in agreement. Differences between the methods result from approximations in the SCF theory and equilibration problems in MC simulations. The two methods are thus complementary.

“Protein-Like” Copolymers: Effect of Polymer Architecture on the Performance in Bioseparation Process

Full Paper: Recently, a new class of copolymers, socalled protein-like copolymers has been predicted theoretically by computer simulation. In these copolymers, the conformation of the copolymer determines the exposure of certain comonomer units to the outer solution. Depending on the conformation, copolymer molecules with essentially the same comonomer composition could have pronouncedly different properties. The authors demonstrated experimentally such behavior in case of poly[(N- vinylcaprolactam)-co-(N-vinylimidazole)] (Dokl. Chem. 2001, 375, 637). One more group of copolymers with protein-like behavior is copolymers of N-isopropylacrylamide with N-vinylimidazole. Poly[(N-isopropylacrylamide)-co-(N-vinylimidazole)] was synthesized by radical polymerization and separated into two fractions using immobilized metal affinity chromatography on Cu 2 + - loaded iminodiacetic acid Sepharose CL 6B (Cu 2 + -IDA- sepharose). The unbound fraction which passed through the column and bound fraction eluted with ethylenediaminetetraacetic acid, disodium salt (EDTA) solution differed significantly in molecular weight, 1.4 × 10 6 and 1.35 × 10 5 , respectively but were very close in comonomer composition, 7.8 and 9.1 mol-% of imidazole, respectively. The composition of bound fraction was confirmed by titration of imidazole groups. Despite close chemical composition, the bound and unbound fraction behaved differently with respect to temperature-induced phase separation at different pH values, the dependence of hydrodynamic diameter on pH and concentration of Cu 2 + - ions, and the coprecipitation of soybean trypsin inhibitor with the copolymer in the presence of Cu 2 + -ions. The differences in the behavior of copolymer fractions are rationalized assuming that the bound fraction presents a protein-like copolymer.

Self-Organization of Quasi-Random Copolymers

Copolymer macromolecules composed of chemi-cally different units are capable of forming partiallyordered spatial structures with diverse morphology [1,2]. The ability to self-assemble is dictated by a differentcharacter of interactions of copolymer units, on the onehand, and by covalent bonding of units within the samemacromolecule, on the other. The latter factor preventsthe separation of the system into homogeneous macro-scopic phases, which can, under definite conditions,stabilize some types of microdomain structures. As arule, a similar phenomenon is treated as microphaseseparation or order–disorder transition. The character-istics of this transition and morphological features ofnascent superstructures are determined by both theexternal parameters and the structure of the copolymeritself. The microphase separation theory mostly dealswith diblock or regular multiblock AB copolymerscomposed of two types of units (A and B) [1, 2]. Thephase behavior of AB copolymers with a random(“quasi-random”) distribution of units along the chainis of particular interest.In this paper, we report on the behavior of a specifictype of quasi-random copolymer in which the distribu-tion of A and B units is similar in a certain sense to theprimary sequence of amino acids in globular proteins.The sequence design scheme for generating such a pro-teinlike copolymer was suggested in [3, 4]. The essenceof this scheme is in chemical modification of the unitslying at the surface of a primary homopolymer globule.Through interaction with a dissolved reagent, some ofthe A units convert into B units. By analogy with globu-lar proteins, these units can be considered to be, respec-tively, hydrophobic and hydrophilic ones. The statisticalproperties of nascent AB sequences are characterized byspecific long-range correlations [5], which, as might beexpected, will influence the phase behavior of similarcopolymers. Such a macromolecule is characterized bythe following parameters: the length N ; the composi-tion, i.e., the fractions of A and B units ϕ

“Protein-Like” Copolymers: Effect of Polymer Architecture on the Performance in Bioseparation Process

Recently, a new class of copolymers, so-called protein-like copolymers has been predicted theoretically by computer simulation. In these copolymers. the conformation of the copolymer determines the exposure of certain comonomer units to the outer solution. Depending on the conformation, copolymer molecules with essentially the same comonomer composition could have pronouncedly different properties. The authors demonstrated experimentally such behavior in case of poly[(N- vinylcaprolactam)-co-(N-vinylimidazole)] (Dokl. Chem. 2001,375, 637). One more group of copolymers with protein-like behavior is copolymers of N-isopropylacryl-amide with N-vinylimidazole. Poly[(N-isopropylacryl-amide)-co-(N-vinylimidazole)] was synthesized by radical polymerization and separated into two fractions using immobilized metal affinity chromatography on Cu2+-loaded iminodiacetic acid Sepbarose CL 6B (Cu2+-IDA-sepharose). The unbound fraction which passed through the column and bound fraction eluted with ethylenediaminetetraacetic acid, disodium salt (EDTA) solution differed significantly in molecular weight, 1.4 x 10(6) and 1.35 x 10(5), respectively but were very close in comonomer composition, 7.8 and 9.1 mol-% of imidazole, respectively. The composition of bound fraction was confirmed by titration of imidazole groups. Despite close chemical composition, the bound and unbound fraction behaved differently with respect to temperature-induced phase separation at different pH values, the dependence of hydrodynamic diameter on pH and concentration of Cu2+- ions, and the coprecipitation of soybean trypsin inhibitor with the copolymer in the presence of Cu2+-ions. The differences in the behavior of copolymer fractions are rationalized assuming that the bound fraction presents a protein-like copolymer. The dependence of hydrodynamic diameter of bound (closed symbols) and unbound (open symbols) poly(NI-PAAM-VI) at different Cu2+/vinylimidazole ratios (n(Ca)/n(VI)), The polymer concentration was 4.5 mg (.) ml(-1) and pH 7.5 was obtained in all systems by using 0.010 m HEPES as a buffer. Mean values from five correlation functions are given. (Less)

Protein-like copolymers: computer simulation

The notion of protein-like AB copolymers is introduced. Such copolymers can be generated with the help of the “ instant image” of a dense homopolymer globule by assigning that the monomeric units closer to the globular surface are of A type, while the core is formed by the B type units. After that the primary structure of the chain is fixed, and one introduces different interaction potentials for A and B units. In doing so, we have in mind mainly aqueous systems and analogy with globular proteins, therefore A units are regarded as hydrophilic, and B units as hydrophobic. By means of Monte Carlo simulation using the bond fluctuation model we study the coil–globule transition for a protein-like copolymer upon the increase of attraction of hydrophobic B units, and compare the results with those for random AB copolymers. From the analysis of the primary structure of protein-like copolymers one can see that the “ degree of blockiness” of the protein-like sequence is higher than for random copolymers, therefore the copolymers with the “ random-block” primary structure are generated for comparison as well (the average length of A and B sequences being the same as for protein-like copolymers). It is shown that the coil–globule transition in protein-like copolymers occurs at higher temperatures, is more abrupt and has faster kinetics than for random copolymers with the same A/ B composition and for random-block copolymers with the same A/ B composition and “ degree of blockiness”. The globules of protein-like copolymers exhibit a dense micelle-like core of hydrophobic B units stabilized by the long dangling loops of hydrophilic A units. Apparently, a protein-like copolymer “ inherits” some of the properties of the “ parent globule” which is reflected in the special long-range correlations in primary structure.