Enantiomers of 3‑Methylspermidine Selectively Modulate Deoxyhypusine Synthesis and Reveal Important Determinants for Spermidine Transport

Eukaryotic translation initiation factor 5A (eIF5A) is essential for cell proliferation, becoming functionally active only after post-translational conversion of a specific Lys to hypusine [N ε-(4-amino-2-hydroxybutyl)­lysine]. Deoxyhypusine synthase (DHS) is the rate-limiting enzyme of this two-ste...

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Published in:ACS chemical biology 2015-06, Vol.10 (6), p.1417-1424
Main Authors: Hyvönen, Mervi T, Khomutov, Maxim, Petit, Marine, Weisell, Janne, Kochetkov, Sergey N, Alhonen, Leena, Vepsäläinen, Jouko, Khomutov, Alex R, Keinänen, Tuomo A
Format: Article
Language:English
Subjects:
Biological Transport
Cell Line, Tumor
Eflornithine - pharmacology
Eukaryotic Translation Initiation Factor 5A
Gene Expression
Humans
Lysine - analogs & derivatives
Lysine - biosynthesis
Lysine - metabolism
Male
Ornithine Decarboxylase Inhibitors - pharmacology
Oxidoreductases Acting on CH-NH Group Donors - antagonists & inhibitors
Oxidoreductases Acting on CH-NH Group Donors - genetics
Oxidoreductases Acting on CH-NH Group Donors - metabolism
Peptide Initiation Factors - genetics
Peptide Initiation Factors - metabolism
Protein Processing, Post-Translational
RNA-Binding Proteins - genetics
RNA-Binding Proteins - metabolism
Spermidine - analogs & derivatives
Spermidine - metabolism
Spermidine - pharmacology
Stereoisomerism
Substrate Specificity
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Summary:Eukaryotic translation initiation factor 5A (eIF5A) is essential for cell proliferation, becoming functionally active only after post-translational conversion of a specific Lys to hypusine [N ε-(4-amino-2-hydroxybutyl)­lysine]. Deoxyhypusine synthase (DHS) is the rate-limiting enzyme of this two-step process, and the polyamine spermidine is the only natural donor of the butylamine group for this reaction, which is very conservedhypusine biosynthesis suffers last when the intracellular spermidine pool is depleted. DHS has a very strict substrate specificity, and only a few spermidine analogs are substrates of the enzyme and can support long-term growth of spermidine-depleted cells. Herein, we compared the biological properties of earlier unknown enantiomers of 3-methylspermidine (3-MeSpd) in deoxyhypusine synthesis, in supporting cell growth and in polyamine transport. Long-term treatment of DU145 cells with α-difluoromethylornithine (inhibitor of polyamine biosynthesis) and (R)-3-MeSpd did not cause depletion of hypusinated eIF5A, and the cells were still able to grow, whereas the combination of α-difluoromethylornithine with a racemate or (S)-3-MeSpd caused cessation of cell growth. Noticeably, DHS preferred the (R)- over the (S)-enantiomer as a substrate. (R)-3-MeSpd competed with [14 C]-labeled spermidine for cellular uptake less efficiently than the (S)-3-MeSpd (K i = 141 μM vs 19 μM, respectively). The cells treated with racemic 3-MeSpd accumulated intracellularly mainly (S)-3-MeSpd, but not DHS substrate (R)-3-MeSpd, explaining the inability of the racemate to support long-term growth. The distinct properties of 3-MeSpd enantiomers can be exploited in designing polyamine uptake inhibitors, facilitating drug delivery and modulating deoxyhypusine synthesis.
ISSN:1554-8929
1554-8937
DOI:10.1021/cb500938e