Human spermidine synthase: cloning and primary structure.
Using a synthetic deoxyoligonucleotide mixture constructed for a tryptic peptide of the bovine enzyme as a probe, cDNA coding for the full-length subunit of spermidine synthase was isolated from a human decidual cDNA library constructed on phage lambda gt11. After subcloning into the Eco RI site of pBR322 and propagation, both strands of the insert were sequenced using a shotgun strategy. Starting from the first start codon, which was immediately preceded by a GC-rich region including four overlapping CCGCC consensus sequences, an open reading frame for a 302-amino-acid polypeptide was resolved. This peptide had an Mr of 33,827, started with methionine, and ended with serine. The identity of the isolated cDNA was confirmed by comparison of the deduced amino acid sequence with resolved sequences of the tryptic peptides of bovine spermidine synthase. The coding strand of the cDNA revealed no special regulatory or ribosome-binding signals within 82 nucleotides preceding the start codon and no polyadenylation signal within 247 nucleotides following the stop codon. The coding region, containing a 13-nucleotide repeat close to the 5' end, was longer than, and very different from, that of the bacterial counterpart. This region seems to be of retroviral origin and shows marked homology with sequences found in a variety of human, mammalian, avian, and viral genes and mRNAs. By computer analysis, the first 200 nucleotides of the 5' end of the coding strand appear able to form a very stable secondary structure with a free energy change of -157.6 kcal/mole.(ABSTRACT TRUNCATED AT 250 WORDS)
Wahlfors
J
J
Department of Biochemistry, University of Kuopio, Finland.
Alhonen
L
L
Kauppinen
L
L
Hyvönen
T
T
Jänne
J
J
Eloranta
T O
TO
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D003656:Decidua / Q000201:enzymology; D005260:Female; D006801:Humans; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011247:Pregnancy; D012333:RNA, Messenger / Q000235:genetics; D011436:Spermidine Synthase / Q000235:genetics*; D014166:Transferases / Q000235:genetics*
D012333:RNA, Messenger; D014166:Transferases; D011436:Spermidine Synthase