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and C8
1
Department of Chemistry and Biochemistry and School of Medicine, University of South Carolina, Columbia, SC 29208
Human C8 is composed of three nonidentical subunits (C8
, C8ß,
and C8
) that are encoded in separate genes. In C8 isolated from
serum, these are arranged as a disulfide-linked C8- dimer that is
noncovalently associated with C8ß. In this study, a recombinant form
of C8- was expressed independently of C8ß in insect cells and
COS-7 cells and was shown to be equivalent to serum-derived C8-
with respect to its ability to combine with C8ß and form functional
C8. Also expressed separately were mutant (mut) forms of C8 and
C8 in which the single interchain disulfide bond was eliminated.
MutC8 exhibited the ability to combine with C8ß and express
hemolytic activity, although at a lower level than human C8. Addition
of purified mutC8 increased this activity, presumably by binding to
mutC8. A possible role for C8 as a retinol binding protein was
also investigated. Absorbance spectroscopy and fluorescence emission
and quenching revealed no specific binding of retinol to mutC8.
Together, these results indicate that 1) the biosynthesis and secretion
of C8- is not dependent on C8ß, which is consistent with in
vivo observations in C8ß-deficient humans; 2) C8 can be
synthesized independently of C8; therefore, protection of C8 from
premature membrane interactions during biosynthetic processing is not a
likely function of C8; 3) C8 enhances but is not required for
expression of C8 activity; and 4) C8 does not bind retinol;
therefore, it cannot function as a retinol transport protein.
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