Molecular characterization of human histone H2BK120 Ubiquitin Ligase UBR7
IR@IICB: CSIR-Indian Institute of Chemical Biology, Kolkata
View Archive Info| Field | Value | |
| Title |
Molecular characterization of human
histone H2BK120 Ubiquitin Ligase UBR7
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| Creator |
Dasgupta, Anirban
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| Subject |
Structural Biology & Bioinformatics
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| Description |
Ubiquitination of histones is involved in the maintenance of chromatin dynamics and genome
stability through its ability to regulate gene expression and DNA damage repair. Nucleosomal
histones H2A and H2B have been predominantly targeted for monoubiquitination. In
mammals, Histone H2B gets mono-ubiquitinated in the conserved Lys-120 (K120),
corresponding to Lys-123 (K123) in Budding Yeast. This initiates a degradation independent
pathway which facilitates recruitment of other ‘writer’ enzymes involved in H3K4 and H3K79
methylation which subsequently helps establish an epigenetic crosstalk network with the
H2BK120Ub mark. In humans, H2B K120 monoubiquitination is associated with disruption
of chromatin compaction and increased transcription. Recently, our work had identified
Ubiquitin Protein Ligase E3 Component N-Recognin 7 (UBR7) to be a novel H2BK120
monoubiquitin ligase which pairs with E2 conjugate UbcH6 for its E3 ligase function. In the
present thesis work, to gain insight into the previously unknown mechanism of UBR7 mediated
ubiquitin transfer, I have extensively mapped the E3-E2 binding interface between UBR7 and
UbcH6. The results obtained suggest that atypical PHD finger of UBR7 is crucial for
interaction with UbcH6 and concomitant substrate histone H2B monoubiquitination. The
critical loop regions of UbcH6 involved in UBR7 interaction were identified and their role in
UBR7 mediated H2B monoubiquitination was assessed. The histone H2B C-terminal tail (114-
125) is necessary and was found sufficient by itself to undergo UBR7/UbcH6-mediated
monoubiquitination. PHD finger was found to mediate dimerization of UBR7 and I used SECMALS
to determine the molecular mass of UBR7-PHD in solution. Furthermore, the residues
mapped to the dimerization interface were found implicated in cancer and were also critical in
regulating E3 ubiquitin ligase function of UBR7. Upon dimer deficiency, the E2 and substrate
binding of UBR7 was found to be compromised. Finally, I have also compared the mode of
ubiquitin transfer of UBR7 to RNF20, a previously reported H2B K120 ubiquitin ligase,
through single turnover ubiquitin discharge assays. Interestingly, unlike RNF20, the
UbcH6~Ub hydrolysis mediated by UBR7 requires substrate histone H2B association.
Substrate H2B binding to UBR7 brings about a change in conformation within the PHD finger
which was found to be critical for efficient ubiquitin transfer. RNF20 was not subjected to any
such conformational change. Thus, the mechanism of ubiquitin transfer by UBR7 was found
significantly distinct from that of RNF20.
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| Date |
2022
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| Type |
Thesis
NonPeerReviewed |
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| Format |
application/pdf
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| Identifier |
http://www.eprints.iicb.res.in/2854/1/Anirban_Dasgupta__Thesis.pdf
Dasgupta, Anirban (2022) Molecular characterization of human histone H2BK120 Ubiquitin Ligase UBR7. PhD thesis, JADAVPUR UNIVERSITY. |
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| Relation |
http://www.eprints.iicb.res.in/2854/
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