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Structural Studies of Hyperthermophilic Viruses
Structural Studies of Hyperthermophilic Viruses. Crenarchaeal
viruses are very unique; they are usually found to exist in extreme thermal
environments (greater than 70°C), and frequently at extreme pH (1.0 < pH < 4.0),
conditions commonly encountered in the hot springs of Yellowstone. Sulfolobus
Spindle Virus 1 (SSV1) and Sulfolobus Turreted Icosahedral
Virus (STIV) are arguably the best studied of these crenarchaeal viruses. The
genomes of both viruses have been sequenced, but the majority of the open reading
frames have yet to be assigned any function. We are engaged in structural
and biochemical studies of the SSV and STIV proteomes in an effort to elucidate
the functions of these proteins.
Due to lack of similarity with the public databases, sequence data alone has
largely failed to unlock the functions of these viral genomes. It is unlikely,
however, that the majority of a genome is unique. It is well known that tertiary
(3D) similarities between proteins persist far longer on the evolutionary time
scale than either primary (amino acid) or genomic sequence (DNA) similarities. Our
work over the last three years clearly demonstrates the validity of this approach,
tertiary structure does suggest function. Importantly, the insights gained
from our structural studies suggest function for an ever increasing number of
viral proteins. These structure-function relationships are relevant not
only to the viruses under study (SSVs and STIV), but for the Crenarchaea in general. Further,
our work has also contributed to structural studies illustrating important
evolutionary relationships among viruses in the three domains of life, and
to an understanding of protein stability at extreme temperatures.
Structural Studies of the Response to Oxidative Stress in Archaea. Oxidative
stress, in which reactive oxygen species (ROS) react indiscriminately with
DNA, proteins and lipids, is a universal phenomenon experienced by organisms
in all domains of life. Cumulative damage from ROS is now thought to
contribute to numerous disease states. Thus, an understanding of the
molecular response to oxidative stress is of significant medical importance.
In collaboration with the Douglas and Young laboratories, we have initiated
studies of the oxidative stress response in Sulfolobus solfataricus,
a model Crenarchaeote. Initial work has identified a DPS-like protein
(SsDPSL) whose transcript levels are highly up regulated in response to hydrogen
peroxide. The structure of SsDPSL reveals a new class of proteins containing
a “thioferritin” motif. Current efforts are focused on elucidating
the functional details of this motif, and studies on similar proteins in human
pathogens. In addition to our studies of the DPSL protein, microarray
data has identified several other proteins that are strongly up regulated in
response to oxidative stress. We are beginning structural characterization
of these proteins as well.
Fig.
1. Surface Residues on One Face of the ORF D-63 Dimer are Highly Conserved.
Conserved positive, negative, and non-charged residues are colored blue, red,
and grey respectively. Non-conserved residues are colored yellow. 180 degree
rotation shows strong sequence conservation between SSV1, SSV2, and SSV RH
along one face perpendicular to the 2-fold symmetry axis. Two smaller regions
of conserved surface are found on the opposite side. ORFD-63 might serve as
an adaptor molecule in the assembly of larger macromolecular complexes vital
to viral propagation.
Fig.
2. The Structure of F-93 Reveals a Winged Helix DNA Binding Motif. The DNA
has been docked here by superposition of the winged helix protein 1F4K and
its accompanying DNA onto F-93, followed by deletion of the 1F4K protein.
This leaves the 1F4K DNA docked to F-93, and suggests the manner in which
F-93 might bind DNA. The structure suggests that F-93 is a transcription
factor regulating expression of viral and/or host proteins.
Current Laboratory Personnel:
George Gauss, Postdoctoral Associate
Eric Larson, Ph.D. Student
Smita Menon, Ph.D. Student.
Annop Sendamarai, Ph.D. Student
Nathanael Lintner, Undergraduate Student
Björn Trankle, Undergraduate Student
Brian Eilers, Technical Support Staff
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