Abstract
Deoxyhypusine synthase (DHS) is a transferase enabling the formation of deoxyhypusine, which is the first, rate-limiting step of a unique post-translational modification: hypusination. DHS catalyses the transfer of a 4-aminobutyl moiety of polyamine spermidine to a specific lysine of eukaryotic translation factor 5A (eIF5A) precursor in a nicotinamide adenine dinucleotide (NAD)-dependent manner. This modification occurs exclusively on one protein, eIF5A, and it is essential for cell proliferation. Malfunctions of the hypusination pathway, including those caused by mutations within the DHS encoding gene, are associated with conditions such as cancer or neurodegeneration. Here, we present a series of high-resolution crystal structures of human DHS. Structures were determined as the apoprotein, as well as ligand-bound states at high-resolutions ranging from 1.41 to 1.69 Å. By solving DHS in complex with its natural substrate spermidine (SPD), we identified the mode of substrate recognition. We also observed that other polyamines, namely spermine (SPM) and putrescine, bind DHS in a similar manner as SPD. Moreover, we performed activity assays showing that SPM could to some extent serve as an alternative DHS substrate. In contrast to previous studies, we demonstrate that no conformational changes occur in the DHS structure upon spermidine-binding. By combining mutagenesis and a light-scattering approach, we show that a conserved “ball-and-chain” motif is indispensable to assembling a functional DHS tetramer. Our study substantially advances our knowledge of the substrate recognition mechanism by DHS and may aid the design of pharmacological compounds for potential applications in cancer therapy.
Highlights
Hypusination is the post-translational modification of lysine to the unusual amino acid hypusine (N-ε-(4-amino-2-hydroxybutyl)lysine) [1]
Hypusination is the most specific post-translational modification known to date [4] and it is essential for eukaryotic translation factor 5A (eIF5A) activity [5]. eIF5A is involved in elongation [6], termination [7] and the stimulation of peptide bond formation [8], and it facilitates protein synthesis by resolving polyproline-induced ribosomal stalling; its role seems indispensable in proline repeat-rich protein synthesis [9]
The structure of apo deoxyhypusine synthase (DHS) was determined by molecular replacement using the Protein Data Bank (PDB) entry 1DHS as a search model and refined to a resolution of 1.52 Å with Rwork/Rfree values of 15.42%/17.62%
Summary
Hypusination is the post-translational modification of lysine to the unusual amino acid hypusine (N-ε-(4-amino-2-hydroxybutyl)lysine) [1]. This modification involves two enzymes, namely deoxyhypusine synthase (DHS) and deoxyhypusine hydroxylase (DOHH). Hypusine is present only in the eukaryotic translation factor 5A (eIF5A) [2,3]. Hypusination is the most specific post-translational modification known to date [4] and it is essential for eIF5A activity [5]. The hypusination of eIF5A is essential for its activity in promoting cell proliferation [3,10,11,12]. There is evidence of eIF5A being important in different diseases such as diabetes, cancer and HIV infection [13,14,15,16,17]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
