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Purification and partial characterization of human polyamine synthases
E O Kajander1, L I Kauppinen, R L Pajula
1Department of Biochemistry, University of Kuopio, Finland.
Abstract:
Spermidine synthase was purified to apparent homogeneity from human spleens (8700-fold) by affinity chromatography. The native enzyme was composed of two subunits of identical Mr (35,000) and showed an apparent Mr of 62,000 in pore-gradient gel electrophoresis. Its pI was 5.1, Spermine synthase was purified to apparent homogeneity from placenta (5300-fold) and from kidney (4600-fold). The native enzyme was composed of two subunits of identical Mr (45,000) and showed an apparent Mr of 78,000 in pore-gradient gel electrophoresis. In isoelectric focusing it revealed two bands, with pI values of 4.9 and 5.0. Both synthases were present in all human tissues studied, but revealed a clear tissue-specific pattern. Mouse antisera against spermidine synthase revealed only one band, of Mr 35,000, in all purified enzyme preparations and in crude human tissue extracts in immunoblotting. Antisera against spermine synthase showed an immunoreactive band corresponding to the Mr of the subunit of spermine synthase. These antisera did not indicate any cross-reactivity in immunoblotting. Thus spermine synthase and spermidine synthase do not share homologous antigenic sites and are totally different proteins.
Insights
Spermidine synthase and spermine synthase are distinct proteins found in human tissues. Immunological analysis confirmed no shared antigenic sites, highlighting their unique structures and functions.
Area of Science:
- Biochemistry
- Enzymology
- Human Physiology
Background:
- Spermidine synthase and spermine synthase are key enzymes in polyamine metabolism.
- Understanding their biochemical properties and tissue distribution is crucial for comprehending polyamine homeostasis.
Purpose of the Study:
- To purify and characterize spermidine synthase and spermine synthase from human tissues.
- To investigate the tissue-specific expression patterns of these enzymes.
- To determine the immunological relationship between spermidine synthase and spermine synthase.
Main Methods:
- Affinity chromatography for enzyme purification.
- Pore-gradient gel electrophoresis and isoelectric focusing for molecular weight and pI determination.
- Immunoblotting using specific antisera to assess antigenic relationships.
Main Results:
- Spermidine synthase purified from spleen showed subunits of Mr 35,000 and pI 5.1.
- Spermine synthase purified from placenta and kidney showed subunits of Mr 45,000 and pI values of 4.9 and 5.0.
- Both enzymes exhibited tissue-specific expression patterns.
- Antisera against each enzyme showed no cross-reactivity, indicating distinct antigenic sites.
Conclusions:
- Spermidine synthase and spermine synthase are immunologically distinct proteins.
- These enzymes possess different subunit molecular weights and isoelectric points.
- Their presence in various human tissues suggests important, potentially tissue-specific roles in polyamine synthesis.