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Related Experiment Videos

Human acid sphingomyelinase.

Stephanie Lansmann1, Christina G Schuette, Oliver Bartelsen

  • 1Kekulé-Institut für Organische Chemie und Biochemie, Universität Bonn, Germany.

European Journal of Biochemistry
|March 13, 2003
PubMed
Summary

This study identifies the disulfide bond pattern of human acid sphingomyelinase (haSMase), crucial for understanding its role in Niemann-Pick disease and cellular signaling. The findings confirm an intramolecular SAP-type activator domain.

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Area of Science:

  • Biochemistry
  • Structural Biology
  • Enzymology

Background:

  • Human acid sphingomyelinase (haSMase) degrades sphingomyelin, and its deficiency causes Niemann-Pick disease.
  • Despite being cloned over a decade ago, complete structural information for haSMase remains limited.
  • Understanding haSMase structure is vital for elucidating its roles in cell signaling and apoptosis.

Purpose of the Study:

  • To determine the disulfide bond pattern of human acid sphingomyelinase (haSMase) for the first time.
  • To validate the predicted intramolecular sphingolipid activator protein (SAP)-type activator domain.

Main Methods:

  • Functional recombinant haSMase was expressed using a baculovirus system and purified.
  • Trypsin digestion followed by MALDI-MS analysis identified four disulfide bonds.

Related Experiment Videos

  • Partial reduction and MALDI-PSD analysis identified two additional disulfide bonds.

    • Main Results:

    • The study identified six disulfide bonds in haSMase: Cys120-Cys131, Cys385-Cys431, Cys584-Cys588, Cys594-Cys607, Cys221-Cys226, and Cys227-Cys250.
    • One disulfide bond (Cys120-Cys131) was located in the N-terminal SAP-homologous domain.
    • The disulfide structure of recombinant haSMase matched that of the native enzyme isolated from human placenta.

    Conclusions:

    • The identified disulfide bond pattern provides essential structural insights into haSMase.
    • The results support the hypothesis of an intramolecular SAP-type activator domain within haSMase.
    • This structural information is critical for understanding haSMase function and Niemann-Pick disease pathogenesis.