Thus, the 3D structure of apo α-La directs the nature of the supramolecular
structures resulting from its assembly with LYS. Hence, as already underlined by
Chiti and Dobson [79], subsequent self-assembly of proteins is highly promoted
by conformational change and/or formation of small oligomers. Behind these
structural considerations, the nature of some other constituents such as small ions
is quite important. Theoretical approaches confirm the main role of molecular
structure as well as of the surrounding environment. To predict the morphology
of cross-assembled particles at a given thermodynamic condition, explicit access to
the effective interactions between the macromolecules and the surrounding solutes
is of crucial importance. Several studies indicate that this can be reached by the
coarse-grained Monte Carlo approach that provides such access through a judicious
choice of the interaction parameters [137].
Recent kinetic investigations showed that the temperature, in the range 20–45
C,
affects only the structural re-organisation of the formed final supramolecular
structures. Whatever the temperature, small homogeneous aggregates form rapidly
when the two proteins are mixed that subsequently grow by collision and fusion, as
evidenced by dynamic and static light scattering [138]. However, the amorphous
aggregates formed at 25
C are rapidly converted to well-defined microspheres once
the temperature is increased above 30
C [138].
Working on this protein system, we demonstrated for the first time the possibility
of spontaneous formation of microspheres between two oppositely charged
proteins, under conditions of charge compensation. The final size of formed spheres
seems to be highly dependent on the initial protein concentration. Microspheres of
about 3–5 μm are classically obtained starting with a total protein concentration
around 0.2–0.5 mM. Decreasing the initial total protein concentration to 0.02 mM
leads to the formation of nanospheres with a diameter of about 100–200 nm. Hence,
the particle size is directly correlated to the initial protein concentration. Quantification of proteins in formed microspheres shows a perfect stoichiometry with α-La:
LYS molar ratio of 1, and the two proteins exhibited a perfect spatial co-localisation
in the microspheres [134]. Moreover, the effect of ionic strength on both the
formation and the stability of the microspheres has been described [12]. An ionic
strength above 100 mM completely inhibits the assembly between the two proteins.
However, once formed, a higher salt concentration is needed to dissociate the
microspheres. In this sense, the divalent ions (calcium, magnesium) are more
effective than monovalent ions in the disassembly of the microspheres. Starting
from these published experimental results, Persson and Lund [139] conducted a
pioneering theoretical study on the self-assembly between these two proteins using
Monte Carlo simulations. For such a simulation study, the two proteins, counterions and salt particles are immersed in a spherical simulation cell containing a
continuum solvent described by the dielectric constant of water (Fig. 6). The
authors suggest that the highly uneven charge distribution on α-La is responsible
for the formation of strongly ordered heterodimers that facilitate the formation of
structured mesoscopic aggregates through electrostatic steering [139]. The process
is described as similar to the interaction between an ion and a dipole; LYS has a
clear preference for interaction to the negative end of α-La, which coincides with
Spontaneous Assembly and Induced Aggregation of Food Proteins
89
structures resulting from its assembly with LYS. Hence, as already underlined by
Chiti and Dobson [79], subsequent self-assembly of proteins is highly promoted
by conformational change and/or formation of small oligomers. Behind these
structural considerations, the nature of some other constituents such as small ions
is quite important. Theoretical approaches confirm the main role of molecular
structure as well as of the surrounding environment. To predict the morphology
of cross-assembled particles at a given thermodynamic condition, explicit access to
the effective interactions between the macromolecules and the surrounding solutes
is of crucial importance. Several studies indicate that this can be reached by the
coarse-grained Monte Carlo approach that provides such access through a judicious
choice of the interaction parameters [137].
Recent kinetic investigations showed that the temperature, in the range 20–45
C,
affects only the structural re-organisation of the formed final supramolecular
structures. Whatever the temperature, small homogeneous aggregates form rapidly
when the two proteins are mixed that subsequently grow by collision and fusion, as
evidenced by dynamic and static light scattering [138]. However, the amorphous
aggregates formed at 25
C are rapidly converted to well-defined microspheres once
the temperature is increased above 30
C [138].
Working on this protein system, we demonstrated for the first time the possibility
of spontaneous formation of microspheres between two oppositely charged
proteins, under conditions of charge compensation. The final size of formed spheres
seems to be highly dependent on the initial protein concentration. Microspheres of
about 3–5 μm are classically obtained starting with a total protein concentration
around 0.2–0.5 mM. Decreasing the initial total protein concentration to 0.02 mM
leads to the formation of nanospheres with a diameter of about 100–200 nm. Hence,
the particle size is directly correlated to the initial protein concentration. Quantification of proteins in formed microspheres shows a perfect stoichiometry with α-La:
LYS molar ratio of 1, and the two proteins exhibited a perfect spatial co-localisation
in the microspheres [134]. Moreover, the effect of ionic strength on both the
formation and the stability of the microspheres has been described [12]. An ionic
strength above 100 mM completely inhibits the assembly between the two proteins.
However, once formed, a higher salt concentration is needed to dissociate the
microspheres. In this sense, the divalent ions (calcium, magnesium) are more
effective than monovalent ions in the disassembly of the microspheres. Starting
from these published experimental results, Persson and Lund [139] conducted a
pioneering theoretical study on the self-assembly between these two proteins using
Monte Carlo simulations. For such a simulation study, the two proteins, counterions and salt particles are immersed in a spherical simulation cell containing a
continuum solvent described by the dielectric constant of water (Fig. 6). The
authors suggest that the highly uneven charge distribution on α-La is responsible
for the formation of strongly ordered heterodimers that facilitate the formation of
structured mesoscopic aggregates through electrostatic steering [139]. The process
is described as similar to the interaction between an ion and a dipole; LYS has a
clear preference for interaction to the negative end of α-La, which coincides with
Spontaneous Assembly and Induced Aggregation of Food Proteins
89