Crystal structure of thermospermine synthase from Medicago truncatula and substrate discriminatory features of plant aminopropyltransferases

Abstract

Polyamines are linear polycationic compounds that play a crucial role in the growth and development of higher plants. One triamine (spermidine, SPD) and two tetraamine isomers (spermine, SPM, and thermospermine, TSPM) are obtained by the transfer of the aminopropyl group from decarboxylated -adenosylmethionine to putrescine and SPD. These reactions are catalyzed by the specialized aminopropyltransferases. In that respect, plants are unique eukaryotes that have independently evolved two enzymes, thermospermine synthase (TSPS), encoded by the gene, and spermine synthase, which produce TSPM and SPM, respectively. In this work, we structurally characterize the gene product, TSPS, from the model legume plant (). Six crystal structures of TSPS - one without ligands and five in complexes with either reaction substrate (SPD), reaction product (TSPM), or one of three cofactor analogs (5'-methylthioadenosine,-adenosylthiopropylamine, and adenosine) - give detailed insights into the biosynthesis of TSPM. Combined with small-angle X-ray scattering data, the crystal structures show that TSPS is a symmetric homotetramer with an interdomain eight-stranded β-barrel. Such an assembly and the presence of a hinge-like feature between N-terminal and C-terminal domains give the protein additional flexibility which potentially improves loading substrates and discarding products after the catalytic event. We also discuss the sequence and structural features around the active site of the plant aminopropyltransferases that distinguish them from each other and determine their characteristic substrate discrimination.

Structural Analysis of Phosphoserine Aminotransferase (Isoform 1) From Arabidopsis thaliana– the Enzyme Involved in the Phosphorylated Pathway of Serine Biosynthesis

“…At PSAT1 was cloned and purified using a modified protocol recently applied for the production of Medicago truncatula N-carbamoylputrescine amidohydrolase ( Sekula et al, 2016 ) and thermospermine synthase ( Sekula and Dauter, 2018 ). Isolation of total RNA from leaves of A. thaliana was performed with RNeasy Plant Mini Kit (Qiagen).…”

Polyamines in Microalgae: Something Borrowed, Something New

“…TSMS is very similar to SMS both in primary sequence and biosynthetic mechanism; both use Spd as an acceptor substrate for the dcSAM aminopropyl group, but the products are different. A possible explanation comes from a recent study, which solved the crystal structure of TSMS from Medicago truncatula (MtTSMS) [101]. First, the dcSAM binding site may be important for the differential enzymatic activity.…”

“…In addition, TSMS has a Trp255 in the middle of the polyamine binding pocket, while most SMS have Ile at the same position. The hydrophobic Trp255 pushes the middle amine group closer to the catalytic Asp178 and, in this way, the aminopropyl and aminobutyl moieties of Spd are well orientated to generate TSpm instead of Spm [101].…”

Spermidine Synthase (SPDS) Undergoes Concerted Structural Rearrangements Upon Ligand Binding – A Case Study of the Two SPDS Isoforms From Arabidopsis thaliana

“…Spermine and thermospermine are produced from SPD by spermine synthase (SPMS) and thermospermine synthase (TSPS), respectively. Although both APTs use the same substrate they present significant differences that predestine them to produce distinct products (Sekula and Dauter, 2018). From the group of plant APTs, only the structures of TSPS from Medicago truncatula ( Mt TSPS, PDB ID 6bq2) (Sekula and Dauter, 2018), and SPDS1 from Arabidopsis thaliana ( At SPDS1, PDB ID 1xj5, Center for Eukaryotic Structural Genomics) are known, however there is no published structural characterization of At SPDS1.…”