The crystal structure of KSHV ORF57 reveals dimeric active sites important for protein stability and function
Authors: Meng-Lu Hu
Ke Lan
Yu-Hui Dong
Xiao-Xi Lin
Zhi-Ming Zheng
Zeng-Qiang Gao
Vladimir Majerciak
Fei Yuan
Lei Bai
Journal: PLoS pathogens (2018)
Discipline: MICROBIOLOGY
Keywords: Kaposi's sarcoma-associated herpesvirus; ORF57; dimerization; stability; viral mRNA
Affiliations: State Key Laboratory of Virology, College of Life Sciences, Medical Research Institute, Wuhan University, Wuhan, P. R. China., Beijing Synchrotron Radiation Facility, Institute of High Energy Physics, Chinese Academy of Sciences, Beijing, China., Tumor Virus RNA Biology Section, RNA Biology Laboratory, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, Maryland, United States of America.

Abstract:  Kaposi's sarcoma-associated herpesvirus (KSHV) is a γ-herpesvirus closely associated with Kaposi's sarcoma, primary effusion lymphoma and multicentric Castleman disease. Open reading frame 57 (ORF57), a viral early protein of KSHV promotes splicing, stability and translation of viral mRNA and is essential for viral lytic replication. Previous studies demonstrated that dimerization of ORF57 stabilizes the protein, which is critical for its function. However, the detailed structural basis of dimerization was not elucidated. In this study, we report the crystal structures of the C-terminal domain (CTD) of ORF57 (ORF57-CTD) in both dimer at 3.5 Å and monomer at 3.0 Å. Both structures reveal that ORF57-CTD binds a single zinc ion through the consensus zinc-binding motif at the bottom of each monomer. In addition, the N-terminal residues 167-222 of ORF57-CTD protrudes a long "arm" and holds the globular domains of the neighboring monomer, while the C-terminal residues 445-454 are locked into the globular domain in cis and the globular domains interact in trans. In vitro crosslinking and nuclear translocation assays showed that either deletion of the "arm" region or substitution of key residues at the globular interface led to severe dimer dissociation. Introduction of point mutation into the zinc-binding motif also led to sharp degradation of KSHV ORF57 and other herpesvirus homologues. These data indicate that the "arm" region, the residues at the globular interface and the zinc-binding motif are all equally important in ORF57 protein dimerization and stability. Consistently, KSHV recombinant virus with the disrupted zinc-binding motif by point mutation exhibited a significant reduction in the RNA level of ORF57 downstream genes ORF59 and K8.1 and infectious virus production. Taken together, this study illustrates the first structure of KSHV ORF57-CTD and provides new insights into the understanding of ORF57 protein dimerization and stability, which would shed light on the potential design of novel therapeutics against KSHV infection and related diseases. 

Retracted: No

PubPeer comments:
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 It is worrying that Konopleva is listed with several clinical trials. , 
 I think that they used the same control for both western blots. It is clearly separated from the other conditions in the experiment. Possibly not very elegant, but still I do not think that it is wrong. , 
 The highlighted panels are paired with different panels in A and C. , 
 Figure 5. Much more similar than you would expect. https: imgur.com CAgwVwZ ,