An N-Terminal Extension to UBA5 Adenylation Domain Boosts UFM1 Activation: Isoform-Specific Differences in Ubiquitin-like Protein Activation. Issue 3 (1st February 2019)

• Record Type: Journal Article
• Authors: Soudah, Nadine; Padala, Prasanth; Hassouna, Fouad; Kumar, Manoj; Mashahreh, Bayan; Lebedev, Andrey A.; Isupov, Michail N.; Cohen-Kfir, Einav; Wiener, Reuven
• Is Part Of: Journal of molecular biology. Volume 431:Issue 3(2019)
• Page Start: 463
• Page End: 478
• Publication Date: 2019-02-01
• Subjects: UFM1 -- UBA5 -- ubiquitin-like proteins -- E1 activating enzymes -- crystal structure; UFM1 ubiquitin fold modifier 1 -- UBA5 ubiquitin-like modifier-activating enzyme 5 -- UFC1 ubiquitin-fold modifier-conjugating enzyme 1 -- UIS UFM1-interacting sequence -- ITC isothermal titration calorimetry -- TSA thermal shift assay -- BMOE bis-maleimidoethane; Molecular biology -- Periodicals; Biology -- Periodicals; Biochemistry -- Periodicals; Bacteriology -- Periodicals; Molecular Biology -- Periodicals; Biochemistry -- Periodicals; Biologie moléculaire -- Périodiques; Biologie -- Périodiques; Biochimie -- Périodiques; Moleculaire biologie; Biochemistry; Biology; Molecular biology; Periodicals; 572.805
• DOI: 10.1016/j.jmb.2018.10.007
• Languages: English
• ISSNs: 0022-2836

Abstract

Abstract: Modification of proteins by the ubiquitin-like protein, UFM1, requires activation of UFM1 by the E1-activating enzyme, UBA5. In humans, UBA5 possesses two isoforms, each comprising an adenylation domain, but only one containing an N-terminal extension. Currently, the role of the N-terminal extension in UFM1 activation is not clear. Here we provide structural and biochemical data on UBA5 N-terminal extension to understand its contribution to UFM1 activation. The crystal structures of the UBA5 long isoform bound to ATP with and without UFM1 show that the N-terminus not only is directly involved in ATP binding but also affects how the adenylation domain interacts with ATP. Surprisingly, in the presence of the N-terminus, UBA5 no longer retains the 1:2 ratio of ATP to UBA5, but rather this becomes a 1:1 ratio. Accordingly, the N-terminus significantly increases the affinity of ATP to UBA5. Finally, the N-terminus, although not directly involved in the E2 binding, stimulates transfer of UFM1 from UBA5 to the E2, UFC1. Graphical Abstract: Highlights: The N-terminal extension of UBA5 long isoform stimulates UFM1 activation. Crystal structures of UBA5 long isoform with and without UFM1 are reported. The N-terminal extension of UBA5 increases UBA5's affinity to ATP. The N-terminal extension of UBA5 contributes to UFM1 transfer to UFC1.