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This vector was used to generate p HSV-HBx-R96E using targeted mutagenesis (Quik Change II XL site-directed mutagenesis kit, Bio connect (Agilent Technologies, Santa Clara, CA, USA)), according to the manufacturer’s instructions.The primer pairs used were HBx-R96E-F: 5′-CCCAAGGTCTTACATAAGGAGACTCTTGGAGTCCCAGC-3′, and HBx-R96E-R: 5′-GCTGGGAGTCCAAGAGTCTCCTTATGTAAGACCTTGGG-3′.DDB1 can form a complex with Cullin-4A (Cul4A) and an E3 ring ubiquitin ligase that ubiquitinates substrate proteins, which are subsequently degraded by the proteasome.

HBx invariably transactivates transcription from circular DNA templates, irrespective of the promoter driving transcription [16,17,18,19].

Therefore, we reason that HEK293 cells may also be suitable for the identification of the putative target, which is degraded upon its interaction with HBx and DDB1.

We systematically analyzed HBx-interacting proteins in HEK 293 cells in the presence and absence of HBx expression.

The capacity to transactivate RNA transcription is related to the in vivo function of HBx [20] and can be used to assess HBx functionality [17].

For its function in vivo, HBx requires an interaction with the damaged DNA binding protein 1 (DDB1) [21,22,23,24,25,26,27].

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