Replication Protein A (RPA) from human cell
s i
s a
stable complexof 70-, 32-, and 14-kDa
subunit
s that i
s required for multiple proce
sse
s in DNA metaboli
sm.RPA bind
s with high affinity to
single-
stranded DNA and interact
s with multiple protein
s, includingprotein
s required for the initiation ofSV40 DNA replication, DNA polymera
se
![](/image<font color=)
s/gifchar
s/alpha.gif" BORDER=0> and SV40 large T antigen.We have u
sed a
serie
s of mutantderivative
s of RPA to map the region
s of RPA required for
specificprotein-protein interaction
s andhave examined the role
s of the
se interaction
s in DNA replication.T antigen, DNA polymera
se
![](/image<font color=)
s/gifchar
s/alpha.gif" BORDER=0> and theactivation domain of VP16 all have overlapping
site
s of interaction inthe N-terminal half (re
sidue
s 1-327)of the 70-kDa
subunit of RPA. In addition, the interaction
sitefor DNA polymera
se
![](/image<font color=)
s/gifchar
s/alpha.gif" BORDER=0> i
s compo
sed oftwo functionally di
stinct region
s, one (re
sidue
s 1-~170) which
stimulate
s polymera
se activity and a
second (re
sidue
s ~170-327) which increa
se
s polymera
se proce
ssivity.In the latter, both the directprotein-protein interaction and
ssDNA-binding activitie
s of RPA wereneeded for RPA to modulatepolymera
se proce
ssivity. We al
so found that SV40 T antigeninhibited the ability of RPA to increa
seproce
ssivity of DNA polymera
se
![](/image<font color=)
s/gifchar
s/alpha.gif" BORDER=0>,
sugge
sting that thi
s activity ofRPA may be important for elongationbut not during the initiation of DNA replication. DNA polymera
se
![](/image<font color=)
s/gifchar
s/alpha.gif" BORDER=0>, but not T antigen al
so interactedwith the 32- and/or 14-kDa
subunit
s of RPA, but the
se interaction
s didnot
seem to effect polymera
seactivity.