Biochemical characterisation and novel classification of monofunctional S-adenosylmethionine decarboxylase of Plasmodium falciparum.

Plasmodium falciparum like other organisms is dependent on polyamines for proliferation. Polyamine biosynthesis in these parasites is regulated by a unique bifunctional S-adenosylmethionine decarboxylase/ornithine decarboxylase (PfAdoMetDC/ODC). Only limited biochemical and structural information is available on the bifunctional enzyme due to the low levels and impurity of an instable recombinantly expressed protein from the native gene. Here we describe the high level expression of stable monofunctional PfAdoMetDC from a codon-harmonised construct, which permitted its biochemical characterisation indicating similar catalytic properties to AdoMetDCs of orthologous parasites. In the absence of structural data, far-UV CD showed that at least on secondary structure level, PfAdoMetDC corresponds well to that of the human protein. The kinetic properties of the monofunctional enzyme were also found to be different from that of PfAdoMetDC/ODC as mainly evidenced by an increased K(m). We deduced that complex formation of PfAdoMetDC and PfODC could enable coordinated modulation of the decarboxylase activities since there is a convergence of their k(cat) and lowering of their K(m). Such coordination results in the aligned production of decarboxylated AdoMet and putrescine for the subsequent synthesis of spermidine. Furthermore, based on the results obtained in this study we propose a new AdoMetDC subclass for plasmodial AdoMetDCs.

Molecular machines encoded by bacterially-derived multi-domain gene fusions that potentially synthesize, N-methylate and transfer long chain polyamines in diatoms.

Silica glass formation in diatoms requires the biosynthesis of unusual, very long chain polyamines (LCPA) composed of iterated aminopropyl units. Diatoms processively synthesize LCPA, N-methylate the amine groups and transfer concatenated, N-dimethylated aminopropyl groups to silaffin proteins. Here I show that diatom genomes possess signal peptide-containing gene fusions of bacterially-derived polyamine biosynthetic enzymes S-adenosylmethionine decarboxylase (AdoMetDC) and an aminopropyltransferase, sometimes fused to a eukaryotic histone N-methyltransferase domain, that potentially synthesize and N-methylate LCPA. Fusions of similar, alternatively configured domains but with a catalytically dead AdoMetDC and in one case a Tudor domain, may N-dimethylate and transfer multiple aminopropyl unit polyamines onto silaffin proteins.

S-Adenosylmethionine decarboxylase from the archaeon Methanococcus jannaschii: identification of a novel family of pyruvoyl enzymes.

Polyamines are present in high concentrations in archaea, yet little is known about their synthesis, except by extrapolation from bacterial and eucaryal systems. S-Adenosylmethionine (AdoMet) decarboxylase, a pyruvoyl group-containing enzyme that is required for spermidine biosynthesis, has been previously identified in eucarya and Escherichia coli. Despite spermidine concentrations in the Methanococcales that are several times higher than in E. coli, no AdoMet decarboxylase gene was recognized in the complete genome sequence of Methanococcus jannaschii. The gene encoding AdoMet decarboxylase in this archaeon is identified herein as a highly diverged homolog of the E. coli speD gene (less than 11% identity). The M. jannaschii enzyme has been expressed in E. coli and purified to homogeneity. Mass spectrometry showed that the enzyme is composed of two subunits of 61 and 63 residues that are derived from a common proenzyme; these proteins associate in an (alphabeta)(2) complex. The pyruvoyl-containing subunit is less than one-half the size of that in previously reported AdoMet decarboxylases, but the holoenzyme has enzymatic activity comparable to that of other AdoMet decarboxylases. The sequence of the M. jannaschii enzyme is a prototype of a class of AdoMet decarboxylases that includes homologs in other archaea and diverse bacteria. The broad phylogenetic distribution of this group suggests that the canonical SpeD-type decarboxylase was derived from an archaeal enzyme within the gamma proteobacterial lineage. Both SpeD-type and archaeal-type enzymes have diverged widely in sequence and size from analogous eucaryal enzymes.