HIV Viral
Infectivity Factor BC-box in complex with ElonginB and ElonginC
David Torres '16 and Joey Duronio '16
Contents:
I. Introduction
The Human Immunodefiency Virus
(HIV) as well as other similar retroviruses contain a viral
infectivity factor (Vif) that inhibits certain anti-viral activity
in human cells. Vif is used in viral replication by hijacking the
Cullin5 E3 ubiquitin ligase, which consists of Cullin5, ElonginBC,
andRbx2, and targeting APOBEC3G for degradation. APOBEC3G is a
cellular enzyme and part of the APOBEC superfamily of proteins. This
family of proteins plays a role in anti-viral immunity. Vif inhibits
APOBEC3G from entering the new budding virus and hypermutating the
genome to render the virus essentially dead.
Vif is able to hijack the
Cullin5 E3 ubiquitin ligase by binding to ElonginBC via the BC-box
domain. It then uses the hosts own ubiquitination and degradation
system to destroy the APOBEC3G.
General
Structure
of Elongin-Cullin-SOCS-box (ECS)
A normal
cellular ECS ubiquitin ligase is made up of ElonginB (EloB), ElonginC
(EloC),
Cullin5 (Cul5), Rbx2, and a SOCS-box protein. SOCS-box is formally
known as the
suppressor of cytokine signaling box proteins. SOCS box proteins are
characterized by a conserved interaction domain, which offers a link
between
cellular substrates and the E3 ubiquitin ligase. This conserved domain
of SOCS
box includes the BC box, which binds EloB and EloC (EloBC), and the
Cullin box,
which is hypothesized to participate in the binding of either Cul5 or
Cul2.
Crystal structures show that the SOCS box has a conserved region
composed of a
BC box helix, a small 90°
turn, a second short helix ending in a region rich with prolines, a
loop, and a
third helix. The first helix is considered as the BC box and the rest
is
referred to as the Cullin box.
Vif
Binding
APOBEC3G
Vif uses
conserved regions to influence the degradation of APOBEC3G. Vif acts
as the
substrate of APOBEC3G by mimicking the SOCS box domain of a normal
cellular E3
ubiquitin ligase. Research proposes that the Vif N-terminal domain
interacts
with APOBEC3G, while its C-terimanl domain recruits the E3 ligase
through two
conserved regions. HIV Vif contains a BC-box region that interacts
with EloC
and is crucial for recruiting the cellular E3 ubiquitin ligase. A
number of Vif
residues V142, L145, L148, A149, A152, AND L153 create the hydrophobic
face
that interacts with EloC. Hydrogen bonding between the backbone
carbonyl of Vif
G143 and EloC Y76 also contributes to the Vif-EloC interaction. A
second E3
ligase-binding domain is a conserved region containing histidine and
cysteine
residues (this region is called the zinc-binding motif). This
zinc-binding
motif interacts with Cul5. Mutations of these His or Cys residues
destroys the
Vif-Cul5 interaction.
The
interaction between a third Vif C-terminal domain, the Vif Cullin box,
and the
E3 ubiquitin ligase has not been thoroughly studied. However, the
Vif-Cul5
interaction has been identified by many experiments. These experiments
were not
able to identify a direct interaction between the Vif Cullin box and
Cul5.
IV. DNA Binding
Once CAP has bound cAMP, it is ready to bind to the DNA.
Binding occurs at the conserved sequence of
5'-AAATGTAGATCACATTT-3'
Hydrogen bonds between the protein and the DNA phsophates occur at the
backbone amide of residue
139, and the side chains of Thr-140,
Ser-179, and Thr-182
In addition to these phosphate interactions, the side chains of Glu-181
and Arg-185, both emanating from the
recognition
helix
directly contact the bases within the major groove of the DNA. Because
of the way that the protein binds to the DNA, a kink of ~40
degrees occurs between nucleotide base pairs six
and seven on each side of the dyad
axis, 5'-TG-3'
This sequence has been shown to favor DNA flexibility and bending in
other systems as well. Because of this kink, an additional five
ionic interactions and four hydrogen bonds are able to occur
between the protein and the DNA strand. Examples of these new
interactions occur between Lys-26, Lys-166,
His-199 and the DNA sugar-phosphate backbone
The DNA bend is integral to the mechanism of transcription activation.
Not only does it place CAP in the proper orientation for
interaction with RNA polymerase, but wrapping the DNA around the
protein may result in direct contacts between upstream DNA and RNA
polymerase.
V. Activating Regions
Transcription activation by CAP requires more than merely
the binding of cAMP and binding and bending of DNA. CAP contains
an "activating region" that has been proposed to participate in
direct protein-protein interactions with RNA polymerase and/or
other basal transcription factors. Specifically, amino acids 156,
158, 159,
and 162
have been proposed to be critical for transcription activation by CAP.
These amino acids are part of a surface loop composed of residues
152-166
Researchers have concluded that the third and final step in
transcription activation is this direct protein-protein contact
between amino acids 156-162 of CAP, and RNA polymerase.
VI. References
Gunasekera, Angelo, Yon W. Ebright, and
Richard H. Ebright. 1992. DNA Sequence Determinants for Binding of
the Escherichia coli Catabolite Gene Activator Protein. The
Journal of Biological Chemistry 267:14713-14720.
Schultz, Steve C., George C. Shields, and
Thomas A. Steitz. 1991. Crystal Structure of a CAP-DNA complex:
The DNA Is Bent by 90 degrees Science 253: 1001-1007.
Vaney, Marie Christine, Gary L. Gilliland,
James G. Harman, Alan Peterkofsky, and Irene T. Weber. 1989.
Crystal Structure of a cAMP-Independent Form of Catabolite Gene
Activator Protein with Adenosine Substituted in One of Two
cAMP-Binding Sites. Biochemistry 28:4568-4574.
Weber, Irene T., Gary L. Gilliland, James
G. Harman, and Alan Peterkofsky. 1987. Crystal Structure of a
Cyclic AMP-independent Mutant of Catabolite Activator Protein. The
Journal of Biological Chemistry 262:5630-5636.
Zhou, Yuhong, Ziaoping Zhang, and Richard
H. Ebright. 1993. Identification of the activating region of
catabolite gene activator protein (CAP): Isolation and
characterization of mutants of CAP specifically defective in
transcription activation. Proceedings of the National
Academy of Sciences of the United States of America
90:6081-6085.
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