6J26

Crystal structure of the branched-chain polyamine synthase from Thermococcus kodakarensis (Tk-BpsA) in complex with N4-bis(aminopropyl)spermidine and 5'-methylthioadenosine

Summary for 6J26

• Entry DOI: 10.2210/pdb6j26/pdb
• Descriptor: N(4)-bis(aminopropyl)spermidine synthase, 5'-DEOXY-5'-METHYLTHIOADENOSINE, N~1~,N~1~,N~1~-tris(3-azaniumylpropyl)butane-1,4-diaminium, ... (5 entities in total)
• Functional Keywords: n(4)-bis(aminopropyl)spermidine synthase, polyamine biosynthesis, spermidine, branched polyamines, transferase
• Biological source: Thermococcus kodakarensis (strain ATCC BAA-918 / JCM 12380 / KOD1)
• Total number of polymer chains: 2
• Total formula weight: 86071.34

Authors

Mizohata, E.,Toyoda, M.,Fujita, J.,Inoue, T. (deposition date: 2018-12-31, release date: 2019-06-26, Last modification date: 2023-11-22)

Primary citation

Hidese, R.,Toyoda, M.,Yoshino, K.I.,Fukuda, W.,Wihardja, G.A.,Kimura, S.,Fujita, J.,Niitsu, M.,Oshima, T.,Imanaka, T.,Mizohata, E.,Fujiwara, S.
The C-terminal flexible region of branched-chain polyamine synthase facilitates substrate specificity and catalysis.
Febs J., 286:3926-3940, 2019

PubMed Abstract: Branched-chain polyamine synthase (BpsA) catalyzes sequential aminopropyl transfer from the donor, decarboxylated S-adenosylmethionine (dcSAM), to the acceptor, linear-chain polyamine, resulting in the production of a quaternary-branched polyamine via tertiary branched polyamine intermediates. Here, we analyzed the catalytic properties and X-ray crystal structure of Tth-BpsA from Thermus thermophilus and compared them with those of Tk-BpsA from Thermococcus kodakarensis, which revealed differences in acceptor substrate specificity and C-terminal structure between these two enzymes. To investigate the role of the C-terminal flexible region in acceptor recognition, a region (QDEEATTY) in Tth-BpsA was replaced with that in Tk-BpsA (YDDEESSTT) to create chimeric Tth-BpsA C9, which showed a severe reduction in catalytic efficiency toward N -aminopropylnorspermidine, but not toward N -aminopropylspermidine, mimicking Tk-BpsA substrate specificity. Tth-BpsA C9 Tyr and Thr contributed to discrimination between tertiary branched-chain polyamine substrates, suggesting that the C-terminal region of BpsA recognizes acceptor substrates. Liquid chromatography-tandem mass spectrometry analysis on a Tk-BpsA reaction mixture with dcSAM revealed two aminopropyl groups bound to two of five aspartate/glutamate residues (Glu , Asp , Asp , Glu , and Glu ) in the C-terminal flexible region. Mutating each of these five amino acid residues to asparagine/glutamine resulted in a slight decrease in activity. The quadruple mutant D342N/D343N/E344Q/E345Q exhibited a severe reduction in catalytic efficiency, suggesting that these aspartate/glutamate residues function to receive aminopropyl chains. In addition, the X-ray crystal structure of the Tk-BpsA ternary complex bound to N -bis(aminopropyl)spermidine revealed that Asp and Glu interacted with the aminopropyl moiety in N -aminopropylspermidine.

PubMed: 31162806
DOI: 10.1111/febs.14949

Experimental method

X-RAY DIFFRACTION (2 Å)