Biochem. J. (2003) 373
(661–667) (Printed in Great Britain)
Accelerated publication
Genomic identification and biochemical characterization of a second spermidine/spermine N1-acetyltransferase
Ying CHEN1, Slavoljub VUJCIC1, Ping LIANG, Paula DIEGELMAN, Debora L. KRAMER and Carl W. PORTER2
Grace Cancer Drug Center and Department of Genetics (P.L.), Roswell Park Cancer Institute, Elm and Carlton Streets, Buffalo, NY 14263, U.S.A.
In the polyamine back-conversion pathway, spermine and spermidine are first acetylated by spermidine/spermine N1-acetyl-transferase (SSAT-1) and then oxidized by polyamine oxidase to produce spermidine and putrescine respectively. Herein we apply homology-search methods to identify novel sequences belonging to a second SSAT, SSAT-2, with a chromosomal location at 17p13.1, which is distinct from SSAT-1 at Xp22. Human SSAT-2 cDNA derived from small-cell lung carcinoma was deduced to encode a 170-amino-acid protein having 46% sequence identity and 64% sequence similarity with SSAT-1. When transiently transfected into HEK-293 cells, SSAT-1 decreased spermidine and spermine pools by 
30%, while, at the same time, significantly increasing putrescine, N1-acetylspermidine, N1-acetylspermine and N1,N12-diacetylspermine pools. By contrast, transfected SSAT-2 had no effect on intracellular polyamine or acetylated polyamine pools. When enzyme activity was assayed on enzyme extracts from transfected cells, both SSAT-1 and SSAT-2 demonstrated much higher acetylating activity than vector-transfected cells. The data suggest that, in intact cells, SSAT-2 may be compartmentalized or it may be inefficient at low intracellular polyamine concentrations. By substituting candidate substrates in the enzyme assay, we determined that SSAT-1 shows the substrate preference norspermidine=spermidine
spermine>N1-acetylspermine>putrescine, whereas SSAT-2 shows the preference norspermidine>spermidine=spermineN1-acetylspermine=putrescine. Analysis of mRNA levels in cell lines and ESTs (expressed sequence tags) from various tissues by digiNorthern (a web-based tool for virtually displaying expression profiles of query genes based on EST sequences) indicated that SSAT-1 tends to be more widely and highly expressed than SSAT-2. While SSAT-1 mRNA was inducible by polyamine analogues in a variety of cell lines, SSAT-2 was not. The existence of an active, but possibly sequestered, SSAT-2 enzyme suggests that, under certain conditions, it may be recruited into basal or perturbed polyamine metabolism.
Key words: genomics, polyamine oxidase, polyamine analogue, spermidine, spermidine/spermine N1-acetyltransferase, spermine oxidase.
Abbreviations used: N1-AcSpd, N1-acetylspermidine; N1-AcSpm, N1-acetylspermine; DASpm, N1,N12-diacetylspermine; DESpm, N1,N12-diethylspermine; DEHSpm, N1,N14-diethylhomospermine; DENSpm, N1,N11-diethylnorspermine; EST, expressed sequence tag; GNAT, generic N-acetyltransferase; norSpd, norspermidine; ODC, ornithine decarboxylase; PAO, polyamine oxidase; Put, putrescine; SMO, spermine oxidase; Spd, spermidine; Spm, spermine; SSAT, spermidine/spermine N1-acetyltransferase.
1Shared first authorship.
2To whom correspondence should be addressed (e-mail carl.porter@roswellpark.org).
Received 20 May 2003/12 June 2003; accepted 13 June 2003
Published as BJ Immediate Publication 13 June 2003, DOI 10.1042/BJ20030734
The Biochemical Society, London ©2003
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Unofficial 2010 Impact Factor for BJ Energy calculated by dividing the number of times articles published in 2008 and 2009 were cited in 2010 (based on a search of the Web of ScienceSM Thomson Reuters) by the number of articles published in 2008 and 2009.
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