SASDC69: Human Rev7 dimer in complex with Rev3 peptide @ 4.6mg/mL

Basic information

Entry

Database: SASBDB / ID: SASDC69

Sample

Human Rev7 dimer in complex with Rev3 peptide @ 4.6mg/mL

• Mitotic spindle assembly checkpoint protein MAD2B (protein), hRev7-WT dimer, Homo sapiens
• DNA polymerase zeta catalytic subunit (protein), Rev3-RBM2, Homo sapiens
• DNA polymerase zeta catalytic subunit (protein), Rev3-RBM2, Homo sapiens

Function / homology

Function:

somatic diversification of immunoglobulins involved in immune response / DNA damage response, signal transduction resulting in transcription / negative regulation of transcription regulatory region DNA binding / zeta DNA polymerase complex / positive regulation of isotype switching / positive regulation of extracellular matrix assembly / negative regulation of transcription by competitive promoter binding / negative regulation of cell-cell adhesion mediated by cadherin / JUN kinase binding / negative regulation of epithelial to mesenchymal transition / positive regulation of double-strand break repair via nonhomologous end joining / negative regulation of ubiquitin protein ligase activity / mitotic spindle assembly checkpoint signaling / telomere maintenance in response to DNA damage / negative regulation of double-strand break repair via homologous recombination / error-prone translesion synthesis / positive regulation of epithelial to mesenchymal transition / Translesion synthesis by REV1 / Translesion synthesis by POLK / Translesion synthesis by POLI / actin filament organization / regulation of cell growth / double-strand break repair via homologous recombination / negative regulation of canonical Wnt signaling pathway / negative regulation of DNA-binding transcription factor activity / negative regulation of protein catabolic process / DNA-templated DNA replication / spindle / double-strand break repair / positive regulation of peptidyl-serine phosphorylation / site of double-strand break / chromosome / 4 iron, 4 sulfur cluster binding / RNA polymerase II-specific DNA-binding transcription factor binding / DNA-directed DNA polymerase / DNA-directed DNA polymerase activity / cell division / nucleotide binding / chromatin / nucleolus / positive regulation of DNA-templated transcription / negative regulation of transcription by RNA polymerase II / DNA binding / nucleoplasm / nucleus / metal ion binding / cytoplasm

Domain/homology:

Domain of unknown function DUF4683 / Domain of unknown function (DUF4683) / DNA polymerase zeta catalytic subunit / C4-type zinc-finger of DNA polymerase delta / C4-type zinc-finger of DNA polymerase delta / Mad2-like / HORMA domain / HORMA domain / HORMA domain profile. / HORMA domain superfamily / DNA polymerase family B, thumb domain / DNA-directed DNA polymerase, family B, multifunctional domain / DNA polymerase family B signature. / DNA-directed DNA polymerase, family B, conserved site / DNA polymerase family B / DNA polymerase family B, exonuclease domain / DNA-directed DNA polymerase, family B, exonuclease domain / DNA polymerase, palm domain superfamily / DNA polymerase type-B family / DNA-directed DNA polymerase, family B / Ribonuclease H superfamily / Ribonuclease H-like superfamily / DNA/RNA polymerase superfamily

Component:

DNA polymerase zeta catalytic subunit / Mitotic spindle assembly checkpoint protein MAD2B

• Biological species: Homo sapiens (human)
• Citation: Journal: Proc Natl Acad Sci U S A / Year: 2018; Title: Rev7 dimerization is important for assembly and function of the Rev1/Polζ translesion synthesis complex.; Authors: Alessandro A Rizzo / Faye-Marie Vassel / Nimrat Chatterjee / Sanjay D'Souza / Yunfeng Li / Bing Hao / Michael T Hemann / Graham C Walker / Dmitry M Korzhnev / ; Abstract: The translesion synthesis (TLS) polymerases Polζ and Rev1 form a complex that enables replication of damaged DNA. The Rev7 subunit of Polζ, which is a multifaceted HORMA (Hop1, Rev7, Mad2) protein with roles in TLS, DNA repair, and cell-cycle control, facilitates assembly of this complex by binding Rev1 and the catalytic subunit of Polζ, Rev3. Rev7 interacts with Rev3 by a mechanism conserved among HORMA proteins, whereby an open-to-closed transition locks the ligand underneath the "safety belt" loop. Dimerization of HORMA proteins promotes binding and release of this ligand, as exemplified by the Rev7 homolog, Mad2. Here, we investigate the dimerization of Rev7 when bound to the two Rev7-binding motifs (RBMs) in Rev3 by combining in vitro analyses of Rev7 structure and interactions with a functional assay in a Rev7 cell line. We demonstrate that Rev7 uses the conventional HORMA dimerization interface both to form a homodimer when tethered by the two RBMs in Rev3 and to heterodimerize with other HORMA domains, Mad2 and p31 Structurally, the Rev7 dimer can bind only one copy of Rev1, revealing an unexpected Rev1/Polζ architecture. In cells, mutation of the Rev7 dimer interface increases sensitivity to DNA damage. These results provide insights into the structure of the Rev1/Polζ TLS assembly and highlight the function of Rev7 homo- and heterodimerization.

Contact author

• Alex Rizzo (University of Connecticut, Storrs, CT, USA)

Models

Sample

• Sample: Name: Human Rev7 dimer in complex with Rev3 peptide @ 4.6mg/mL; Specimen concentration: 4.6 mg/ml / Entity id: 857 / 858 / 859
• Buffer: Name: 20 mM HEPES, 10 mM DTT, 5% glycerol / pH: 8

Entity #857

Name: hRev7-WT dimer / Type: protein
Description: Mitotic spindle assembly checkpoint protein MAD2B
Formula weight: 24.391 / Num. of mol.: 2 / Source: Homo sapiens / References: UniProt: Q9UI95
Sequence:

GMTTLTRQDL NFGQVVADVL CEFLEVAVHL ILYVREVYPV GIFQKRKKYN VPVQMSCHPE LNQYIQDTLH CVKPLLEKND VEKVVVVILD KEHRPVEKFV FEITQPPLLS ISSDSLLSHV EQLLRAFILK ISVCDAVLDH NPPGCTFTVL VHTREAATRN MEKIQVIKDF PWILADEQDV HMHDPRLIPL KTMTSDILKM QLYVEERAHK GS

Entity #858

Name: Rev3-RBM2 / Type: protein / Description: DNA polymerase zeta catalytic subunit / Formula weight: 3.273 / Num. of mol.: 1 / Source: Homo sapiens / References: UniProt: O60673
Sequence:

MEDKKIVIMP CKCAPSRQLV QVWLQAKE

Entity #859

Name: Rev3-RBM2 / Type: protein / Description: DNA polymerase zeta catalytic subunit / Formula weight: 3.273 / Num. of mol.: 1 / Source: Homo sapiens / References: UniProt: O60673
Sequence:

MEDKKIVIMP CKCAPSRQLV QVWLQAKE

Experimental information

• Beam: Instrument name: Cornell High Energy Synchrotron Source (CHESS) G1; City: Ithaca, NY / 国: USA / Type of source: X-ray synchrotron / Wavelength: 0.1267 Å
• Detector: Name: Finger Lakes CCD / Type: CCD

Scan

Title: Human Rev7 dimer in complex with Rev3 peptide @ 4.6mg/mL
Measurement date: May 14, 2016 / Storage temperature: 4 °C / Cell temperature: 4 °C / Exposure time: 1 sec. / Number of frames: 10 / Unit: 1/A /

	Min	Max
Q	0.0086	0.2729

Result

Type of curve: single_conc
Comments: Based on the affinity of the Rev7 dimer, at this concentration (and below) there is a mixture of monomer and dimer in the solution. This dataset was not used for modeling

	Experimental	Standard	Standard error
MW	49.7 kDa	49.7 kDa	0.06

	Guinier	Guinier error
Forward scattering, I0	0.145	0.0018
Radius of gyration, Rg	2.91 nm	0.006

	Min	Max
Guinier point	1	63