1SWT

CORE-STREPTAVIDIN MUTANT D128A IN COMPLEX WITH BIOTIN AT PH 4.5


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.00 Å
  • R-Value Free: 0.308 
  • R-Value Observed: 0.209 

Starting Model: experimental
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This is version 1.4 of the entry. See complete history


Literature

A structural snapshot of an intermediate on the streptavidin-biotin dissociation pathway.

Freitag, S.Chu, V.Penzotti, J.E.Klumb, L.A.To, R.Hyre, D.Le Trong, I.Lybrand, T.P.Stenkamp, R.E.Stayton, P.S.

(1999) Proc Natl Acad Sci U S A 96: 8384-8389

  • DOI: https://doi.org/10.1073/pnas.96.15.8384
  • Primary Citation of Related Structures:  
    1SWS, 1SWT

  • PubMed Abstract: 

    It is currently unclear whether small molecules dissociate from a protein binding site along a defined pathway or through a collection of dissociation pathways. We report herein a joint crystallographic, computational, and biophysical study that suggests the Asp-128 --> Ala (D128A) streptavidin mutant closely mimics an intermediate on a well-defined dissociation pathway. Asp-128 is hydrogen bonded to a ureido nitrogen of biotin and also networks with the important aromatic binding contacts Trp-92 and Trp-108. The Asn-23 hydrogen bond to the ureido oxygen of biotin is lengthened to 3.8 A in the D128A structure, and a water molecule has moved into the pocket to replace the missing carboxylate interaction. These alterations are accompanied by the coupled movement of biotin, the flexible binding loop containing Ser-45, and the loop containing the Ser-27 hydrogen bonding contact. This structure closely parallels a key intermediate observed in a potential of mean force-simulated dissociation pathway of native streptavidin, where the Asn-23 hydrogen bond breaks first, accompanied by the replacement of the Asp-128 hydrogen bond by an entering water molecule. Furthermore, both biotin and the flexible loop move in a concerted conformational change that closely approximates the D128A structural changes. The activation and thermodynamic parameters for the D128A mutant were measured and are consistent with an intermediate that has traversed the early portion of the dissociation reaction coordinate through endothermic bond breaking and concomitant gain in configurational entropy. These composite results suggest that the D128A mutant provides a structural "snapshot" of an early intermediate on a relatively well-defined dissociation pathway for biotin.


  • Organizational Affiliation

    Department of Bioengineering, University of Washington, Seattle, WA 98195, USA.


Macromolecules
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Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
PROTEIN (STREPTAVIDIN)
A, B
127Streptomyces avidiniiMutation(s): 1 
UniProt
Find proteins for P22629 (Streptomyces avidinii)
Explore P22629 
Go to UniProtKB:  P22629
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP22629
Sequence Annotations
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  • Reference Sequence
Small Molecules
Binding Affinity Annotations 
IDSourceBinding Affinity
BTN BindingDB:  1SWT Kd: 0 (nM) from 1 assay(s)
ΔH: min: -1.23e+2, max: -6.69e+1 (kJ/mol) from 12 assay(s)
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.00 Å
  • R-Value Free: 0.308 
  • R-Value Observed: 0.209 
  • Space Group: P 21 21 2
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 55.7α = 90
b = 84.8β = 90
c = 50γ = 90
Software Package:
Software NamePurpose
AMoREphasing
SHELXL-97refinement
SADIEdata reduction
SAINTdata reduction
SADABSdata scaling
XPREPdata reduction

Structure Validation

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Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 1999-07-30
    Type: Initial release
  • Version 1.1: 2008-04-27
    Changes: Version format compliance
  • Version 1.2: 2011-07-13
    Changes: Derived calculations, Version format compliance
  • Version 1.3: 2021-11-03
    Changes: Database references, Derived calculations
  • Version 1.4: 2023-08-23
    Changes: Data collection, Refinement description