Activities of Topoisomerase
I in Its Complex with
SRSF1
- Publication date
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Abstract
Human DNA topoisomerase I (topo I) catalyzes DNA relaxation
and
phosphorylates SRSF1. Whereas the structure of topo I complexed with
DNA has been resolved, the structure of topo I in the complex with
SRSF1 and structural determinants of topo I activities in this complex
are not known. The main obstacle to resolving the structure is a contribution
of unfolded domains of topo I and SRSF1 in formation of the complex.
To overcome this difficulty, we employed a three-step strategy: identifying
the interaction regions, modeling the complex, and validating the
model with biochemical methods. The binding sites in both topo I and
SRSF1 are localized in the structured regions as well as in the unfolded
domains. One observes cooperation between the binding sites in topo
I but not in SRSF1. Our results indicate two features of the unfolded
RS domain of SRSF1 containing phosphorylated residues that are critical
for the kinase activity of topo I: its spatial arrangement relative
to topo I and the organization of its sequence. The efficiency of
phosphorylation of SRSF1 depends on the length and flexibility of
the spacer between the two RRM domains that uniquely determine an
arrangement of the RS domain relative to topo I. The spacer also influences
inhibition of DNA nicking, a prerequisite for DNA relaxation. To be
phosphorylated, the RS domain has to include a short sequence recognized
by topo I. A lack of this sequence in the mutants of SRSF1 or its
spatial inaccessibility in SRSF9 makes them inadequate as topo I/kinase
substrates