4EPJ

Crystal Structure of inactive single chain wild-type HIV-1 Protease in Complex with the substrate p2-NC


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.69 Å
  • R-Value Free: 0.227 
  • R-Value Work: 0.198 
  • R-Value Observed: 0.199 

wwPDB Validation   3D Report Full Report


This is version 1.4 of the entry. See complete history


Literature

Structural, kinetic, and thermodynamic studies of specificity designed HIV-1 protease.

Alvizo, O.Mittal, S.Mayo, S.L.Schiffer, C.A.

(2012) Protein Sci 21: 1029-1041

  • DOI: https://doi.org/10.1002/pro.2086
  • Primary Citation of Related Structures:  
    4EP2, 4EP3, 4EPJ, 4EQ0, 4EQJ

  • PubMed Abstract: 

    HIV-1 protease recognizes and cleaves more than 12 different substrates leading to viral maturation. While these substrates share no conserved motif, they are specifically selected for and cleaved by protease during viral life cycle. Drug resistant mutations evolve within the protease that compromise inhibitor binding but allow the continued recognition of all these substrates. While the substrate envelope defines a general shape for substrate recognition, successfully predicting the determinants of substrate binding specificity would provide additional insights into the mechanism of altered molecular recognition in resistant proteases. We designed a variant of HIV protease with altered specificity using positive computational design methods and validated the design using X-ray crystallography and enzyme biochemistry. The engineered variant, Pr3 (A28S/D30F/G48R), was designed to preferentially bind to one out of three of HIV protease's natural substrates; RT-RH over p2-NC and CA-p2. In kinetic assays, RT-RH binding specificity for Pr3 increased threefold compared to the wild-type (WT), which was further confirmed by isothermal titration calorimetry. Crystal structures of WT protease and the designed variant in complex with RT-RH, CA-p2, and p2-NC were determined. Structural analysis of the designed complexes revealed that one of the engineered substitutions (G48R) potentially stabilized heterogeneous flap conformations, thereby facilitating alternate modes of substrate binding. Our results demonstrate that while substrate specificity could be engineered in HIV protease, the structural pliability of protease restricted the propagation of interactions as predicted. These results offer new insights into the plasticity and structural determinants of substrate binding specificity of the HIV-1 protease.


  • Organizational Affiliation

    Division of Biology, Biochemistry and Molecular Biophysics Option, California Institute of Technology, Pasadena, California 91125, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
protease, tethered dimer203HIV-1 M:B_ARV2/SF2Mutation(s): 8 
Gene Names: gag-pol
EC: 3.4.23.16
UniProt
Find proteins for P03369 (Human immunodeficiency virus type 1 group M subtype B (isolate ARV2/SF2))
Explore P03369 
Go to UniProtKB:  P03369
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP03369
Sequence Annotations
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  • Reference Sequence

Find similar proteins by:  Sequence   |   3D Structure  

Entity ID: 2
MoleculeChains Sequence LengthOrganismDetailsImage
substrate p2-NCB [auth D]8Human immunodeficiency virus 1Mutation(s): 0 
UniProt
Find proteins for Q9YP46 (Human immunodeficiency virus 1)
Explore Q9YP46 
Go to UniProtKB:  Q9YP46
Entity Groups  
UniProt GroupQ9YP46
Sequence Annotations
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  • Reference Sequence
Small Molecules
Ligands 5 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
GOL
Query on GOL

Download Ideal Coordinates CCD File 
C [auth A],
F [auth A]
GLYCEROL
C3 H8 O3
PEDCQBHIVMGVHV-UHFFFAOYSA-N
BME
Query on BME

Download Ideal Coordinates CCD File 
H [auth A]BETA-MERCAPTOETHANOL
C2 H6 O S
DGVVWUTYPXICAM-UHFFFAOYSA-N
DMS
Query on DMS

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E [auth A]DIMETHYL SULFOXIDE
C2 H6 O S
IAZDPXIOMUYVGZ-UHFFFAOYSA-N
EDO
Query on EDO

Download Ideal Coordinates CCD File 
D [auth A]1,2-ETHANEDIOL
C2 H6 O2
LYCAIKOWRPUZTN-UHFFFAOYSA-N
ACT
Query on ACT

Download Ideal Coordinates CCD File 
G [auth A]ACETATE ION
C2 H3 O2
QTBSBXVTEAMEQO-UHFFFAOYSA-M
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.69 Å
  • R-Value Free: 0.227 
  • R-Value Work: 0.198 
  • R-Value Observed: 0.199 
  • Space Group: P 21 21 21
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 51.001α = 90
b = 58.816β = 90
c = 61.794γ = 90
Software Package:
Software NamePurpose
DENZOdata reduction
SCALEPACKdata scaling
REFMACrefinement
PDB_EXTRACTdata extraction

Structure Validation

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Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2012-06-06
    Type: Initial release
  • Version 1.1: 2013-01-09
    Changes: Database references
  • Version 1.2: 2017-07-26
    Changes: Advisory, Refinement description, Source and taxonomy
  • Version 1.3: 2017-11-15
    Changes: Refinement description
  • Version 1.4: 2024-02-28
    Changes: Advisory, Data collection, Database references, Derived calculations