UC Riverside Undergraduate Research Journal

Computational Prediction of Association Free Energies for the C3d-CR2 Complex and Comparison to Experimental Data
Alexander S. Cheung
Introduction
The immune system of higher vertebrates is made
up of both innate and adaptive immunity [1]. The innate
immune system functions primarily through the activity of
leukocytes that indiscriminately work to rid the body of
any foreign pathogenic substances. Activated by the innate
immune system, the adaptive immune system is principally
made up of memory B and T cells. These cells not only
combat pathogenic threats, but also have the ability to
remember specific pathogens and thus mobilize more
rapidly and launch stronger attacks against the pathogen
each time it is again encountered. By efficiently working
together in concert, the innate and adaptive immune
systems protect the body against disease.
The complement system is a key component of
innate immunity but also serves as a link between innate
and adaptive immunities [2]. The complement system
is activated through a complex cascade of catalytic
reactions which involve cleavage of complement proteins
into fragments and formation of protein complexes. The
association of the d-fragment of complement component
C3 (C3d), a globular serum protein, with complement
receptor 2 (CR2), a modular B or T cell surface receptor,
is a crucial link between the innate and adaptive immune
systems. CR2 consists of 15 modules, called complement
control protein (CCP) modules, of which only the first two
modules, CCP1 and CCP2, are known to interact with C3d
([3] and references therein). In this paper we investigate
the interaction between C3d and the first two modules of
CR2, herein referred to as CR2.
There are numerous studies in literature that implicate
charge and electrostatics as having a role in the interaction
between C3d and CR2 [3-10]. It has been established that
electrostatics are essential for many biological functions,
including protein-ligand interactions [3-5], protein stability
[4,11], catalysis [12], conformational transitions [13], and
protein ionization [3-5,11-14]. In previous studies, the
Morikis group has proposed that electrostatics drive C3d-
CR2 recognition [3-5] and used the ionization properties
of C3d and CR2 to demonstrate a correlation between
ionization free energy differences, and association data
from experimental mutagenesis studies [5]. Based on these
studies, a two-step model for association was proposed,
with the first step being recognition, and the second,
binding [3-5,14]. According to this model, recognition
is driven solely by long-range electrostatic interactions
whereas binding involves long- and short/medium-range
electrostatic interactions (including hydrogen bonds, salt
bridges, and van der Waals forces) as well as short-range
hydrophobic interactions and entropic effects. Recognition
is responsible for the formation of a weak, nonspecific
encounter complex, whereas binding is responsible for
the formation of a strong and specific final complex.
Conventional thinking dictates that only mutations at the
binding interface should affect binding abilities. However,
the study by Zhang et al [5] demonstrated that mutations
remote from the association interface can also affect binding
abilities (evaluated as ionization free energy differences),
explaining controversial experimental data that previously
seemed contradictory. This effect of distant mutations on
association is possible only if recognition, as an initial step
in association, involves electrostatic attraction between
protein macrodipoles to form the encounter complex.
Thus, the results of the study performed by Zhang et al [5]
validated the two-step model for C3d-CR2 association.
The goals of our study are to examine whether
similar correlations exist between electrostatic free energies
of association and relative binding ability as well as to
evaluate the effect of solvation on the electrostatic free
energies of association and examine whether correlations
exist between solvation free energy differences and relative
binding ability. We analyzed the effect of the same C3d
and CR2 mutations as in the study by Zhang et al [5] on
C3d-CR2 association. These are mutations for which there
are published experimental data for C3d [9] and CR2 [6].
Figure 1. Molecular representation of C3d-CR2 demonstrating the
topology of the mutated amino acids. Basic and acidic amino acids
are shown in blue and red, respectively. Glu116 was used as the
contact amino acid in association interface and is colored in green.
14 UCR Un d e r g r a d u a t e Re s e a r c h Jo u r n a l