2F9P

Crystal Structure of the Recombinant Human Alpha I Tryptase Mutant D216G in Complex with Leupeptin


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.30 Å
  • R-Value Free: 0.259 
  • R-Value Work: 0.217 
  • R-Value Observed: 0.217 

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This is version 2.2 of the entry. See complete history


Literature

X-ray Structures of Free and Leupeptin-complexed Human alpha I-Tryptase Mutants: Indication for an alpha to beta-Tryptase Transition

Rohr, K.B.Selwood, T.Marquardt, U.Huber, R.Schechter, N.M.Bode, W.Than, M.E.

(2005) J Mol Biol 357: 195-209

  • DOI: https://doi.org/10.1016/j.jmb.2005.12.037
  • Primary Citation of Related Structures:  
    2F9N, 2F9O, 2F9P

  • PubMed Abstract: 

    Tryptases alpha and beta are trypsin-like serine proteinases expressed in large amounts by mast cells. Beta-tryptase is a tetramer that has enzymatic activity, but requires heparin binding to maintain functional and structural stability, whereas alpha-tryptase has little, if any, enzymatic activity but is a stable tetramer in the absence of heparin. As shown previously, these differences can be mainly attributed to the different conformations of the 214-220 segment. Interestingly, the replacement of Asp216 by Gly, which is present in beta-tryptase, results in enzymatically active but less stable alpha-tryptase mutants. We have solved the crystal structures of both the single (D216G) and the double (K192Q/D216G) mutant forms of recombinant human alphaI-tryptase in complex with the peptide inhibitor leupeptin, as well as the structure of the non-inhibited single mutant. The inhibited mutants exhibited an open functional substrate binding site, while in the absence of an inhibitor, the open (beta-tryptase-like) and the closed (alpha-tryptase-like) conformations were present simultaneously. This shows that both forms are in a two-state equilibrium, which is influenced by the residues in the vicinity of the active site and by inhibitor/substrate binding. Novel insights regarding the observed stability differences as well as a potential proteolytic activity of wild-type alpha-tryptase, which may possess a cryptic active site, are discussed.


  • Organizational Affiliation

    Max-Planck-Institut für Biochemie, Abteilung Strukturforschung, Am Klopferspitz 18, 82152 Martinsried, Germany.


Macromolecules
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Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
Tryptase alpha-1A,
C [auth B],
E [auth C],
G [auth D]
245Homo sapiensMutation(s): 2 
Gene Names: TPSAB1TPS1
EC: 3.4.21.59
UniProt & NIH Common Fund Data Resources
Find proteins for Q15661 (Homo sapiens)
Explore Q15661 
Go to UniProtKB:  Q15661
PHAROS:  Q15661
GTEx:  ENSG00000172236 
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupQ15661
Sequence Annotations
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  • Reference Sequence

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Entity ID: 2
MoleculeChains Sequence LengthOrganismDetailsImage
LeupeptinB [auth E],
D [auth F],
F [auth G],
H
4Streptomyces roseusMutation(s): 0 
Sequence Annotations
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  • Reference Sequence
Oligosaccharides

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Entity ID: 3
MoleculeChains Length2D Diagram Glycosylation3D Interactions
alpha-L-fucopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-glucopyranose
I
2N-Glycosylation
Glycosylation Resources
GlyTouCan:  G80587NA
GlyCosmos:  G80587NA
GlyGen:  G80587NA
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 2.30 Å
  • R-Value Free: 0.259 
  • R-Value Work: 0.217 
  • R-Value Observed: 0.217 
  • Space Group: P 21 21 21
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 83.3α = 90
b = 88.23β = 90
c = 162.87γ = 90
Software Package:
Software NamePurpose
DENZOdata reduction
SCALEPACKdata scaling
AMoREphasing
CNSrefinement

Structure Validation

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Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2006-01-31
    Type: Initial release
  • Version 1.1: 2008-05-01
    Changes: Version format compliance
  • Version 1.2: 2011-07-13
    Changes: Atomic model, Database references, Derived calculations, Non-polymer description, Structure summary, Version format compliance
  • Version 1.3: 2012-12-12
    Changes: Other
  • Version 2.0: 2020-07-29
    Type: Remediation
    Reason: Carbohydrate remediation
    Changes: Advisory, Atomic model, Data collection, Derived calculations, Structure summary
  • Version 2.1: 2021-10-20
    Changes: Advisory, Database references, Structure summary
  • Version 2.2: 2023-08-30
    Changes: Data collection, Refinement description