Date of Award


Document Type

Thesis - Pacific Access Restricted

Degree Name

Master of Science (M.S.)



First Advisor

James W. Blanker[unintelligible]


The naturally occuring polyamines play an essential role in cell growth and proliferation.

N8-Acetylspermidine is excreted in rat urine in significant levels (75 nmole/24 hours) but tissue levels of this compound are very low (1.0 nmole/gram), close to the limits of detection. This may be due to rapid degradation of N8-acetylspermidine by an enzyme in the cytoplasm of virtually all rat tissues.

This enzyme N8-acetylspermidine deacetylase catalyzes the deacetylation of N8-acetylpermidine to spermidine and acetic acid.

In order to study the physiologicalf unction of N8-acetylspermidine, we need a means of altering tissue levels of this compound, and inhibition of this deacetylation reaction could produce such an alteration.

In our study, we have used an in vitro deacetylase assay to determine the types of compounds which can inhibit this reaction. The Km for this reaction was 10.5 microM.

Studies on the structural requirements for the inhibitory activity indicated that: (1) increasing the size of the acetamido group leads to a decrease in inhibitory activity, (2) separation of the two primary amine groups with (CH2)8 produces the best inhibitory activity, and (3) removal of the nitrogen in the 4-position decreases the inhibitory activity slightly but increases the stability with respect to a competing enzyme, polyamine oxidase.

Studies on the nature of the active site indicated that: (1) a sulfhydryl group is apparently essential for optimal activity as shown by inhibition with N-ethylmaleimide and also by dithiothreitol and 5,5'-dithio-bis-(nitro-benzoic acid), (2) a seryl hydroxyl group does not appear to be an essential part of the active site since diisopropyl-fluorophosphate has a relatively high Ki. (However, the organophosphate echothiophate is a very effective inhibitor. It has a quartarnary nitrogen which may orient this molecule to its site of action.), and (3) a divalent metal ion may be involved in the enzyme activity as is shown by inhibition with the chelating agents, EDTA and diethylmalonate.



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