1. Loyola Marymount University
Higher Structural Differences in Amino Acid Sequence of gp120 V3 Region in Patients With Rapid CD4+ T Cell Decline Versus Patients with High CD4+ T Cell Variability
Kevin Paiz-Ramirez
Janelle N. Ruiz
Biology
Department of Biology
Loyola Marymount University
March 20, 2010

2. Outline
Background on structure of HIV-1, core gp120, and gp120 complex with CD4 and CD4 co-receptors
Are there structural differences in V3 region of HIV-1 gp120 between participants with rapid CD4+ T cell decline versus patients with high CD4+ T Cell variability?
Results: comparing our groups
Diversity and Divergence of gp120 V3 region structures
Secondary structural predictions of amino sequences
Location and nature of major a.a. changes
Predicted post-translational modifications
Interpretations in Light of Recent Studies
Discussion and Further Studies

3. Entry of HIV Involves a Sequential Interaction in the Structure
The entry of HIV involves specific interactions
The envelope glycoprotein (gp120)
CD4 glycoprotein
CD4+ co-receptors

4. Structure of gp120 Core
Kwong, et, al 1998

5. Importance of core gp120 structure and ternary complex to HIV-1 entry into host cell
Kwong, et al. explored the structure of gp120 in complex with CD4 receptor.
Gp120 is crucial for fusion of HIV-1 to cell surface
Positioning
Timing
CD4 binding induced conformational change in gp120

6. Previous investigation revealed differences in diversity and divergence in DNA sequence of gp120 V3 variants
Patients with high CD4 decline had greater diversity and divergence in DNA sequence of gp120 V3 variants than patients with high variability in CD4 count
Are there structural differences in HIV-1 amino acid sequences between participants with a rapid CD4+ T cell decline versus patients with high CD4+ T Cell variability?
Does this translate to differences in protein sequence and structure?
Linear Progressors
High Variability

7. Methodology for Determining Structural Differences in Sequence between Linear and High Variability Groups
1. Multiple amino acid sequence alignments from first four visits of each patient
2. Determined consensus sequence and sequence(s) with most differences from consensus sequence
3. Predicted secondary structures of sequences using PSIPRED
4. Performed Multiple Sequence alignments for sequences
5. Investigated sites of possible post-translational modifications using PROSITE

8. High Variability group has fewer sequences and less divergence in sequences: Subject 12
Multiple Sequence Alignments
See less amino acid sequences in total and less major amino acid changes between sequences (purple) in High Variability group that Linear progressors
Tables: Represent Major Amino Acid Differences between Consensus Sequence and Sequence with most differences b/w Consensus Sequence 8

9. High Variability group has fewer sequences and less divergence in sequences: Subject 8

10. High Variability group has fewer sequences and less divergence in sequences: Subject 6

11. Linear Progressors have greater number of sequences and greater divergence in sequences: Subject 4
Many more sequences and many more purple major amino acid changes, shown in chart 11

12. High Variability group have greater number of sequences and greater divergence in sequences: Subject 10

13. Structure-Based Sequence Alignment of Core Gp120
Gp120 sequence from Kwong et al.

14. Predicted Secondary Structures of the gp120 V3 Region shows six beta sheets and one alpha helix

15. High Variability shows six beta sheets and one alpha helix
Subject 6
Subject 8
Subject 12

16. Linear Progressors show six beta sheets and one or two alpha helices
Subject 4
Subject 10

17. High Variability Participants Have Greater Major Amino Acid Changes on Protein Surface
Binding Site
*Black 1: Coil region, nonpolar, hydrophobic  Polar Basic
*Black 2: Coil Region, Polar BasicPolar, unchargednonpolar, hydrophobic
*Black 3: Helix 2, Nonpolar, hydorphobicPolar Basic
*Black 4: Coil Region, Polar, unchargednonpolar, hydorophobic
Key:
Blue: Single, fully conserved residues
Green: Conservation of strong groups
Purple: Conservation of weak groups
Black: No consensus

18. Linear Progressor Participants Have Greater Major Amino Acid Changes on Protein Binding Sites
Surface
*Black 1: Beta Sheet 11, polar uncharged nonpolar, hydrophobic
*Black 2: Coil Region, Polar basic nonpolar, hydrophobic
*Black 3: Helix 2, polar uncharged nonpolar, hydrophobic
*Black 4: Coil Region, Polar BasicPolar, uncharged
1st black coil  N-binding site
2nd black coil on Betasheet
3rd black coil On the surface not on a binding site
4th black coil binding site
5th black coil on the surface not on a binding sit
Key:
Blue: Single, fully conserved residues
Green: Conservation of strong groups
Purple: Conservation of weak groups
Black: No consensus

19. Linear Progressor Participants Have Greater Major Amino Acid Changes on Protein Binding Sites
*Black 1: Coil Region, nonpolar, hydrophobic & Polar Basic  Polar Acidic
*Black 2: Beta Sheet 11, Nonpolar Basic  Polar Uncharged
*Black 3: Coil Region, Polar basic nonpolar, hydrophobic
*Black 4: Coil Region, Polar, uncharged  Polar Basic
*Black 5: Coil Region, random
*Black 6: Helix 2, Polar, uncharged & Polar Basic  nonpolar, hydrophobic
*Black 7: Helix 2, nonpolar, hydrophobic & Polar Basic  Polar, uncharged & Polar Acidic
*Black 9: Coil Region, Polar Basic  Polar Uncharged
*Black 10: Coil Region, random
Key:
Blue: Single, fully conserved residues
Green: Conservation of strong groups
Purple: Conservation of weak groups
Black: No consensus

20. Sites of Post Translational Modification: Linear Progressors Have Slightly Greater Post Translational Modifications than High Variability

21. Discussion
Linear progressors had greater number of amino acid sequences (diversity) and greater amount of major amino acid changes (divergence)
This could indicate greater structural differences in linear progressors
Linear Progressors had greater number of major amino acid changes in CD4-binding regions
Linear Progressors showed greater post-translational modifications
Further studies could examine protein structural differences variants within gp41 region

22. Further Investigations Exploring Interactive Regions in gp120
Pancera, et al, examined the gp41 interactive region.
Gp120 elements proximal to gp41 completed a 7 stranded Beta sandwich
Invarient in conformation

23. Further Investigations Exploring Interactive Regions in gp120
Crystal structure was consistent presenting shape changing spacer
Facilitated movement between outer domain and gp41
Provided conformational diversity
Immune evasion

24. References
Kwong PD, Wyatt R, Robinson J, Sweet RW, Sodroski J, and Hendrickson WA. Structure of glycoprotein in complex with the CD4 receptor and a neutralizing human antibody. Nature 1998 Jun 18; 398(6686)
Pancera M, Majeed S, Ban YE, Chen L, Huang CC, Kong L, Kown YD, Stuckey J, Zhou T, Robinson JE, Schief WR, Sodroski J, Wyatt R, and Kwong PD. Structure of HIV-1 gp120 with gp41-interactive region reveals layered envelope architecture and basis of conformational mobility. Proc Natl Acad Sci USA 2010 doi: