Heparin-sepharose as a tool in the subcellular fractionation of a polyamine-rich organ (rat ventral prostate)

Abstract

Heparin-sepharose forms complexes with putrescine, spermidine, and spermine and indirect measurements of its affinity for polyamines gives values similar to those obtained with free heparin. A direct measurement of the binding of heparin-sepharose to spermine gives an apparent dissociation constant (Kd) of 1.5 X 10(-6)M spermine. Unlike free heparin, heparin-sepharose does not cause either disruption of the nuclei or more sutble modifications able to modify their sedimentation behavior. The heparin-sepharose polyamine complex formed by the addition of heparin-sepharose to the homogenate can easily be removed and the homogenate can be processed according to normal schedules. Heparin-sepharose is able to sequester 85% of the exchangeable spermine present in the homogenate of rat ventral prostate. The distribution of the marker enzyme galactosyltransferase (Golgi apparatus) on a sucrose density gradient was followed to assess the usefulness of heparin-sepharose in minimizing the aggregation of cellular organelles brought about by polyamines.