Human Complement Protein C8: The “Hole” Story
Journal of the South Carolina Academy of Science, [
Human Complement Protein C8: The "Hole" Story
James M Sodetz 0
0 Department of Chemistry and Biochemistry, University of South Carolina , Columbia SC 29208 , USA
As the recipient of the 2011 SC Governor's Award for Excellence in Scientific Research, I've been asked to describe my research and some of the accomplishments of my laboratory while at the University of South Carolina. My research is focused on understanding the structure and function of the pore-forming proteins of the human"complement system", and in particular complement protein "C8". The complement system is a group of blood proteins that have a key role in immune defense. Much of what is known today about the structure and function of human C8 can be attributed to work performed over many years by my graduate students and postdoctoral fellows.
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are all noncovalent.
Individually, the five MAC components circulate in blood
as hydrophilic proteins, but when combined they form an
amphiphilic complex capable of intercalating into cell
membranes. The MAC does not degrade membrane lipid but
instead produces a disruptive rearrangement that causes
osmotic lysis of simple cells such as erythrocytes, initiates
intracellular signaling events in nucleated cells, and disrupts
the outer membrane of bacteria. Our own cells contain CD59,
a membrane-anchored protein that protects us from
complement-mediated damage by preventing assembly of a
functional MAC.
The MAC components interact in a highly specific and
sequential manner once C5b is formed. As each intermediate
complex is formed, binding specificity changes and is directed
towards the next component incorporated. Once associated,
the affinity between components is high despite the
noncovalent nature of their interactions. Dissociation can only
be accomplished by solubilization of the membrane and
denaturation of the MAC. The goal of our program has been
to identify structural features that specify the order in which
these proteins bind, and the mechanism by which they
undergo a hydrophilic to amphiphilic transition that enables
them to bind to lipid. Our strategy has been to focus on
identifying structure-function relationships in one component,
namely human C8, and thereby gain insight into how the other
MAC components function. Understanding how the MAC is
assembled and its function may lead to the development of
therapeutic pore-forming analogues that could be used to
attack undesirable human cells, e.g. cancer cells.
The MAC Family Proteins
Human C5b contains two disulfide-linked subunits and is
the largest MAC protein (180 kDa). C6, C7 and C9 are single
chain proteins of approximately 105 kDa, 92 kDa and 72 kDa,
respectively. C8 (151 kDa) is the most complex in that it is
composed of three nonidentical, genetically distinct subunits
(α = 64 kDa; β = 64 kDa and γ = 22 kDa) (
7
). These subunits
are arranged as a disulfide-linked C8α-γ heterodimer that is
noncovalently associated with C8β. The affinity between
C8αγ and C8β is high. Dissociation can only be achieved using
denaturing agents or high ionic strength buffers.
Our discovery in the 1980's that each C8 subunit was
encoded in a different gene was significant because C8 was
thought to be a single gene product, i.e. synthesized as a
single-chain precursor that was posttranslationally cleaved.
This discovery came as we were characterizing human liver
cDNA clones to determine the amino acid sequence of C8 (
8
).
The results established that C8 was composed of three
different proteins rather than one. Our efforts to determine the
sequences and genomic organization of C8 together with the
work of others provided the first evidence of a structural and
evolutionary relationship between the MAC proteins.
Human C6, C7, C8α, C8β, and C9 are homologous and
together form the "MAC family" of proteins. Family members
exhibit sequence similarity and have a highly conserved
modular organization (Fig. 2) (
4,9
). Their genomic structures
are also similar with respect to exon length and boundaries. A
distinctive feature of each is the presence of cysteine-rich
Nand C-terminal modules that are ~40–80 amino acids in
length. The modules exhibit sequence similarity to those
found in a variety of proteins unrelated to complement.
Although not considered a module, the central portion of each
protein (~ 40 kDa) is designated MACPF to emphasize its
conservation among the MAC proteins and its sequence
similarity to perforin, a 70 kDa pore-forming protein released
from secretory granules of cytotoxic T lymphocytes.
The conserved organization of the MAC proteins initially
suggested the modules themselves may mediate
proteinprotein interactions during MAC assembly. This was
consistent with the view that the MAC can be thought of as a
"heteropolymer" formed from structurally similar but distinct
monomeric units. Also noteworthy was the fact these modules
mediate protein-protein interactions in other systems (...truncated)