posted on 2022-10-14, 00:13authored byKarima El Hauadi, Leonor Resina, David Zanuy, Teresa Esteves, Frederico Castelo Ferreira, Maria M. Pérez-Madrigal, Carlos Alemán
CRENKA [Cys-Arg-(NMe)Glu-Lys-Ala,
where
(NMe)Glu refers to N-methyl-Glu],
an anti-cancer pentapeptide that induces prostate tumor necrosis and
significant reduction in tumor growth, was engineered to increase
the resistance to endogenous proteases of its parent peptide, CREKA
(Cys-Arg-Glu-Lys-Ala). Considering their high tendency to aggregate,
the self-assembly of CRENKA and CREKA into well-defined
and ordered structures has been examined as a function of peptide
concentration and pH. Spectroscopic studies and atomistic molecular
dynamics simulations reveal significant differences between the secondary
structures of CREKA and CRENKA. Thus, the restrictions
imposed by the (NMe)Glu residue reduce the conformational
variability of CRENKA with respect to CREKA, which significantly
affects the formation of well-defined and ordered self-assembly morphologies.
Aggregates with poorly defined morphology are obtained from solutions
with low and moderate CREKA concentrations at pH 4, whereas well-defined
dendritic microstructures with fractal geometry are obtained from
CRENKA solutions with similar peptide concentrations at
pH 4 and 7. The formation of dendritic structures is proposed to follow
a two-step mechanism: (1) pseudo-spherical particles are pre-nucleated
through a diffusion-limited aggregation process, pre-defining the
dendritic geometry, and (2) such pre-nucleated structures coalesce
by incorporating conformationally restrained CRENKA molecules
from the solution to their surfaces, forming a continuous dendritic
structure. Instead, no regular assembly is obtained from solutions
with high peptide concentrations, as their dynamics is dominated by
strong repulsive peptide–peptide electrostatic interactions,
and from solutions at pH 10, in which the total peptide charge is
zero. Overall, results demonstrate that dendritic structures are only
obtained when the molecular charge of CRENKA, which is
controlled through the pH, favors kinetics over thermodynamics during
the self-assembly process.