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Orriss, G.

Publications and source records attributed to Orriss, G..

2 recordsLinked to original sources

Cryo-EM structure of the FtsH periplasmic domain reveals functional dynamics

FtsH, an essential AAA+ metalloprotease, maintains cellular homeostasis by degrading misfolded and membrane-associated proteins. Here, we report cryo-EM structures of the Escherichia coli FtsH periplasmic domain (FtsH-PD) revealing insights into its conformational flexibility. Initial 2D class averages suggested three distinct orientations, right-handed and left-handed maps of FtsH-PD, and a map with a different conformation. The 4.9 [A] structure of FtsH-PD exhibits the conserved +{beta} fold, while the 7.3 [A] map with the different conformation displays a 20{o} clockwise rotation of two alpha helices. These findings support a model where conformational changes are present not only in the FtsH cytosolic domain, but also in the periplasmic domain and potentially facilitate substrate translocation through a combination of mechanisms involving both the FtsH-PD and the HflKC complexed with FtsH, along with lipid-scramblase activity to assist in membrane protein extraction. This study points out novel perspectives on how conformational changes in the periplasmic domain contribute to FtsH substrate degradation mechanisms.

biochemistry↗

Dynamic RNA binding and unfolding by nonsense-mediated mRNA decay factor UPF2

Nonsense-mediated mRNA decay (NMD) is an mRNA surveillance pathway involved in translational control and gene expression regulation. Core NMD factors UPF1, UPF2 and UPF3B are conserved from yeast to humans and essential to target mcRNAs with a premature stop codon for decay. UPF2 binding to UPF1 activates UPF1s ATPase and helicase activities, and UPF2 binding to UPF3B is important for its association with the exon-junction complex and efficient NMD. However, UPF2s association with RNA remains largely uncharacterized. Here, we analyze nucleic acid binding, identifying the first and third MIF4G domains of UPF2 as main RNA-/DNA-binding modules. We find that UPF2s MIF4G domain-3 has RNA annealing activity while full-length UPF2 unfolds our reporter hairpin-RNA structure. We show that UPF2 preferentially binds and stabilizes single-stranded RNA (ss-RNA) in a sequence-independent manner. Concomitant to ss-RNA binding, UPF2 undergoes a distinct conformational change in its otherwise highly dynamic structure. UPF2s RNA binding and unfolding activity may support UPF1s helicase and mRNP remodeling activity and, in combination with UPF3B, stabilize UPF1s association with nonsense mRNA.

biochemistry↗