N1-Aminopropylagmatine, a New Polyamine Produced as a Key Intermediate in Polyamine Biosynthesis of an Extreme Thermophile, Thermus thermophilus

Abstract

In the extreme thermophile Thermus thermophilus, a disruption mutant of a gene homologous to speB (coding for agmatinase ‫؍‬ agmatine ureohydrolase) accumulated N 1 -aminopropylagmatine (N 8 -amidino-1,8-diamino-4-azaoctane, N 8 -amidinospermidine), a new compound, whereas all other polyamines produced by the wild-type strain were absent from the cells. Double disruption of speB and speE (polyamine aminopropyltransferase) resulted in the disappearance of N 1 -aminopropylagmatine and the accumulation of agmatine. These results suggested the following. 1) N 1 -Aminopropylagmatine is produced from agmatine by the action of an enzyme coded by speE. 2) N 1 -Aminopropylagmatine is a metabolic intermediate in the biosynthesis of unique polyamines found in the thermophile. 3) N 1 -Aminopropylagmatine is a substrate of the SpeB homolog. They further suggest a new biosynthetic pathway in T. thermophilus, by which polyamines are formed from agmatine via N 1 -aminopropylagmatine. To confirm our speculation, we purified the expression product of the speB homolog and confirmed that the enzyme hydrolyzes N 1 -aminopropylagmatine to spermidine but does not act on agmatine.Polyamines play important roles in cell proliferation and cell differentiation. Common polyamines such as putrescine, spermidine, and spermine are distributed ubiquitously in cells and tissues at relatively high concentrations (1, 2).Thermus thermophilus, of which the genome project was completed using two strains, HB8 and HB27 (Structural-Biological Whole Cell Project at www.srg.harima.riken.go.jp/ thermus/j_index.htm and see Ref. 3, respectively), produces a variety of polyamines including unusually long polyamines and branched ones (4) (see Fig. 9C). These long and branched polyamines have a marked effect of protecting and stabilizing nucleic acids (5, 6) and of activating cell-free polypeptide synthesis at high temperature (7-9).In many organisms, such as bacteria, yeast, animals, and plants, the first step of polyamine biosynthesis is production of putrescine by decarboxylation of L-ornithine (see Fig. 9A). An additional or alternative pathway of putrescine biosynthesis that is often seen in plants and sometimes in bacteria is decarboxylation of L-arginine followed by hydrolysis of agmatine. Agmatine ureohydrolase or agmatinase, coded by the speB gene, catalyzes this second reaction. The next step is production of spermidine and spermine by the addition of an aminopropyl group to putrescine and spermidine, respectively. This reaction is catalyzed by spermidine or spermine synthase (putrescine/spermidine aminopropyltransferase) coded by the speE gene (1).To investigate the polyamine biosynthetic pathway in T. thermophilus, we constructed a disruption strain of the speB gene homolog of T. thermophilus. Disruption of the speB gene homolog resulted in drastic reduction of triamines, longer and branched polyamines without accumulation of agmatine, and in accumulation of an unknown compound. Double disruption of speB and speE gene homologs resulted in disappear...

The First Agmatine/Cadaverine Aminopropyl Transferase: Biochemical and Structural Characterization of an Enzyme Involved in Polyamine Biosynthesis in the Hyperthermophilic Archaeon Pyrococcus furiosus

“…The small amounts of APC and the much higher levels of spermidine measured in P. furiosus cells seem to be in contrast with the activity of ACAPT towards cadaverine that is an order of magnitude higher than that towards putrescine and agmatine observed in vitro (Table 3). Therefore, a detailed kinetic characterization of ACAPT was performed in order to assess whether P. furiosus is able to synthesize spermidine from agmatine using the novel biosynthetic pathway described in the thermophilic bacterium T. thermophilus (33). The K m and V max values for 1,3-diaminopropane, putrescine, agmatine, and cadaverine were measured in the presence of saturating concentrations of decarboxyAdoMet (Fig.…”

“…A novel polyamine biosynthetic pathway is present in T. thermophilus in which an APT utilizes agmatine as a substrate to form N 1 -aminopropylagmatine. This is hydrolyzed by a ureohydrolase, forming spermidine (33).…”

Polyamines in Eukaryotes, Bacteria, and Archaea

“…3) that uses carboxyspermidine dehydrogenase (CASDH) and carboxyspermidine decarboxylase (CASDC) (64,68,69). A variant AdoMet-dependent pathway is present in some bacteria such as the extreme hyperthermophile Thermus thermophilus, where Agm is aminopropylated to form aminopropylagmatine, the substrate for an AUH homologue that then produces Spd (70). There is inherent biosynthetic flexibility in both alternative pathways.…”

Binding and inhibition of human spermidine synthase by decarboxylated S‐adenosylhomocysteine

“…Some aminopropyltransferases such as human spermidine synthase (SpdS) (which acts upon putrescine) and spermine synthase (SpmS) (acting on spermidine) are highly specific for their amine acceptors, [12][13][14] while others, such as those from acute thermophiles, which contain a variety of polyamines not found in mammals, are less discriminating. 12,13,[15][16][17] There are now numerous published structures for aminopropyltransferases including those for SpdS from Thermotoga maritima (TmSpdS), 16 Caenorhabditis elegans, 18 Plasmodium falciparum (PfSpdS), 19 Helicobacter pylori, 20 human (hSpdS), 13 Arabidopsis thaliana (PDB code 2Q41), and Trypanosoma cruzi (PDB code 3BWC), aminopropylagmatine/aminopropylcadaverine synthases from Thermus thermophilus (PDB code 1UIR), Pyrococcus horikoshii (PDB code 2ZSU) and Pyrococcus furiosus, 15 and SpmS from humans (hSpmS). 14 A general mechanism for aminopropyl transfer has been proposed based on kinetic studies of hSpdS, TmSpdS, and hSpmS, their structures with bound substrates and inhibitors, and the results of site-directed mutagenesis of key residues (Fig.…”