Poster abstracts

Poster number 142 submitted by Andrew Sugarman

Structural Dynamics of HIV ESS2p RNA Element

Andrew Sugarman (Chemistry, Case Western Reserve University), Liang-Yuan Chiu (Chemistry, Case Western Reserve University), NaShea Kendrick (Chemistry, Oglethorpe University), William Ford (Chemistry, Case Western Reserve University), Alexander Hansen (CCIC NMR Facility, The Ohio State University), Blanton Tolbert (Chemistry, Case Western Reserve University)

Abstract:
ESS2p is an exonic splicing silencer significant for its role in HIV replication and the virus’s ability
to produce the vital Tat protein. The 43 nucleotide RNA molecule is located downstream of the 3’
splice site A3 on the HIV-1 transcript, where it is typically known to interact with hnRNP H.
Recruitment of this protein is necessary for inhibition of the spliceosome1, and the structure of
ESS2p is responsible for its binding interaction with hnRNP H. In this investigation we report the
solution structure of ESS2p and an assessment of its binding interactions with hnRNP H, and another
vital protein hnRNP A1. NMR techniques including NOESY and RDC experiments were conducted
to evaluate hydrogen bonding, base pairing, and sugar pucker data for ESS2p. Small Angle X-ray
Scattering (SAXS) was also performed to provide a molecular density map of the molecule. Data
from these experiments were combined in XPLOR and AMBER, where a series of molecular
dynamics simulations were carried out to obtain a robust solution structure. In addition, we report
the results of ITC experiments that reveal ESS2p is capable of strong binding interactions with
hnRNP A1 through the apical loop AG sequence. On the other hand, the G-tract sequence of ESS2p
is buried in the base-paired region, as a result, hnRNP H did not show canonical binding profile in
the ITC experiment, indicating that in vivo, hnRNP H likely needs need assistance of other proteins
to unwind the G-tract base pair region of ESS2p to allow recognition.

Keywords: Dynamics, NMR , Simulations