A Mutational Investigation of an HIV Patient

1
A Mutational Investigation of an HIV Patients
GP120 Glycoprotein and its Implications on CD4
Binding Salita Kaistha Usrinus College,
Collegeville PA 19426 November 28, 2003
DNA Results Continued
Structural Results
Background
One advantage that HIV has over our immune
system is its rapid rate of mutation. There are
both synonymous and non-synonymous mutations.
Synonymous mutations are those which involve
changes in the nucleotide sequence, but still
produce the same amino acid sequence.
Non-synonymous mutations are those which change
the nucleotide sequence and the amino acid
sequence. Clone 3 from Visit 1 is what I
consider to be the original strain. All other
clones are variations of this strain. Clone 4
from Visit 4 is an example of a synonymous
mutation, for there is a change in the DNA
sequence (1 nucleotide), but the resulting amino
acid sequence is the same (Fig. 1). Visit 1
clone 1 is an example of a non-synonymous
mutation. There was a change in one nucleotide
of the DNA sequence, however this time the DNA
sequence produces a different amino acid sequence
(Fig. 1).
HIV is a deadly virus killing more than 3
million people a year. This fact alone motivates
researchers to learn as much as possible about
the virus, its structure, and its entry into
cells, in hopes of eventually developing a cure.
Viewed under a microscope an HIV particle would
look like this
Amino Acid Results
Figure 4. This chart presents the 96 amino acid
sequences for the envelope gene of the GP120
glycoprotein of HIV in the various clones of
patient 11 at visits 1 and 4. Amino acids at
positions were subject to mutation at any time
during the two years are highlighted in blue.
Amino acids there were mutated are highlighted in
pink. The amino acids highlighted in green
represent a region on the V1/V2 steam that is
critical for GP120 to bind to CD4.
An HIV particle is approximately 0.0001
mm. An HIV particle consists of two main parts,
the inner core and the viral membrane. This
viral membrane contains two main glycoprotein's,
GP120 and GP41. GP120 allows the HIV particle to
bind to CDR T cells of our immune system. GP41
facilitates membrane fusion.
Figure 5. This is a schematic representation of
a portion of GP120 that interacts with CD4. It
shows the V1/V2 stem on HIVs GP120, which is
encoded for by patient 11s DNA sequence. The
green box highlights 6 amino acids on the V1/V2
stem that are necessary for binding to CD4. The
pink box highlights the fragment of CD4 that the
V1/V2 stem interacts with.
Due to GP120s involvement in cell
recognition and binding, it is of great interest
to researchers. GP120 is encoded for by 500
amino acids, but only a small portion of these
are involved in binding to CD4. In fact, GP120
interactions come from 6 fragments located on the
V1/V2 stem, Loop D, B15-alpha15 excursion,
B20-B21 hairpin, B23, and the B24-alpha5
connection. This coincides with the general
principle that much of a proteins secondary
structure is simply needed to provide a scaffold,
and it is the loops that allow the protein to
function. The progression of HIV is monitored by
an individuals CD4 T cell count.
These sequences demonstrate the fact that
these mutations are not occurring throughout the
entire sequence, but rather at specific portions
of the amino acid sequence over the 2 ½ years
(Fig. 4). In other words certain portions are
subject to mutations, where as other portions are
conserved and experience no mutation over the two
year period. As mentioned earlier, GP120
interacts with CD4 through 6 different fragments
or portions on GP120. The DNA sequences obtained
for patient 11 encode for one of these 6
fragments, that is the V1/V2 stem (Fig. 5). I
have located the 6 amino acids that are critical
for this interaction. They are AHCNVS. Due to
the fact the HIV must bind to CD4 in order to
infect an individuals cells, these 6 amino acids
are critical to the viruss infection and
survival. Thus, logically, these 6 amino acids
would be conserved, and this is precisely what
was discovered. This crucial conserved sequence,
AHCNVS, is highlighted in green in Figure 4.
Prior Research
This study was conducted using research
and data collected by Markham et al. Markham et
al. followed 15 patients for anywhere from 1 ½ -
4 ½ years. They monitored the number of clones
of HIV within the individual and the individuals
CD4 T cell count. The number of clones per visit
ranged anywhere between 2 and 18. The
researchers gathered over 666 DNA sequences.
Figure 2. The amino acids found at selected
positions for all of the clones found at all
visits for patient 11. The selected positions
are those which experienced a mutation at any
time during the two years. The mutated amino
acids are highlighted according to the following
color scheme yellow for Glycines, green for
hydrophobic, pink for negatively charged, blue
for positively charged and purple for polar amino
acids.
References
Methods and Materials
Figure 2 portrays only the amino acid
positions that encountered changes over the 2
years. There are 3 predominant changes taking
place. These changes include converting the
charge ( ? -), converting to a smaller amino
acid (a.a.? G), or a conservative change ( ? ).

Kwong, P., Wyatt, R., and J. Robinson. (1998)
Structure of an HIV gp120 envelope glycoprotein
in complex with the CD4 receptor and a
neutralizing human antibody. Nature 393,
648-659. Markham, R., Wnag, W. and A. Weisstein.
(1998) Patterns of HIV-1 evolution in
individuals with differing rates of CD4 T cell
decline. Proc. Natl. Acad. Sci. 95, 12568-12573.
The Protein Data Bank. (2003)
http//www.rcsb.org/pdb/cgi/explore.cgi?jobgraphi
cspdbId1G9Mpagepid151221070341180. The
Molecules of HIV. (2003) http//www.mcld.co.uk/hi
v/?qHIV20virus20particle. Aids in the World.
(2003) http//www.yale.edu/yaw/world.html.
This study was conducted using the data
gathered by Markham et al. The study focused on
patient 11, a rapid progressor for HIV. Data
was collected for the last two years of the
patients life. Resources used include those
found at PDB, NCBI and ExPASy.
DNA Results
Figure 3. This chart highlights the information
found in the amino acid sequences listed above
and also gives the CD4 T cell counts for the
patient at each visit.
As the disease progresses there is a
simultaneous drop in CD4 T cell counts and an
increase in the number of mutations. Over two
years there are 32 strains and only 17 amino acid
positions altered over the sequence of 96 amino
acids (less than 18). The percentage of
non-synonymous mutations increases as the disease
progresses. There are never more than 3 amino
acid positions altered per strain of HIV,
regardless of the visit number. The number of
strains with an increasing number of mutations
per strain increases as the disease progresses.
Acknowledgements
I would like to thank Dr. Roberts, Tom
Seegar, Derese Getnet and Drew Foy for all their
help in completing this poster.
Figure 1. Codons encoding amino acid positions
that were mutated over the 2 years are
highlighted in blue. The codons containing the
point mutations are highlighted in pink.