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Mixed Macromolecular Crowding: A Protein and Solvent Perspective
journal contribution
posted on 2018-04-19, 08:13 authored by Saikat Biswas, Jayanta Kundu, Sanjib K. Mukherjee, Pramit Kumar ChowdhuryIn the living cell, biomolecules
perform their respective functions
in the presence of not only one type of macromolecules but rather
in the presence of various macromolecules with different shapes and
sizes. In this study, we have investigated the effects of five single
macromolecular crowding agents, Dextran 6, Dextran 40, Dextran 70,
Ficoll 70, and PEG 8000 and their binary mixtures on the modulation
in the domain separation of human serum albumin using a Förster
resonance energy transfer-based approach and the translational mobility
of a small fluorescent probe fluorescein isothiocyanate (FITC) using
fluorescence correlation spectroscopy (FCS). Our observations suggest
that mixed crowding induces greater cooperativity in the domain movement
as compared to the components of the mixtures. Thermodynamic analyses
of the same provide evidence of crossovers from enthalpy-based interactions
to effects dominated by hard-sphere potential. When compared with
those obtained for individual crowders, both domain movements and
FITC diffusion studies show significant deviations from ideality,
with an ideal solution being considered to be that arising from the
sum of the contributions of those obtained in the presence of individual
crowding agents. Considering the fact that domain movements are local
(on the order of a few angstroms) in nature while translational movements
span much larger lengthscales, our results imply that the observed
deviation from simple additivity exists at several possible levels
or lengthscales in such mixtures. Moreover, the nature and the type
of deviation not only depend on the identities of the components of
the crowder mixtures but are also influenced by the particular face
of the serum protein (either the domain I–II or the domain
II–III face) that the crowders interact with, thus providing
further insights into the possible existence of microheterogeneities
in such solutions.