PDBsum entry 4zq9

DNA binding protein/DNA

PDB id: 4zq9

Contents

Protein chains

185 a.a.

DNA/RNA

Metals

_MN ×2

Waters ×58

PDB id:

4zq9

Name:

DNA binding protein/DNA

Title:

X-ray structure of aav-2 obd bound to aavs1 site 3:1

Structure:

Protein rep68. Chain: a, b, c. Fragment: origin binding domain (unp residues 1-206). Engineered: yes. Mutation: yes. DNA (5'- d( Gp Cp Gp Cp Tp Cp Gp Cp Tp Cp Gp Cp Tp Cp Gp Cp Tp Gp Gp Gp C)- 3'). Chain: d.

Source:

Adeno-associated virus 2 (isolate srivastava/1982). Aav-2. Organism_taxid: 648242. Strain: isolate srivastava/1982. Gene: rep68. Expressed in: escherichia coli 'bl21-gold(de3)plyss ag'. Expression_system_taxid: 866768. Synthetic: yes.

Resolution:

2.60Å R-factor: 0.206 R-free: 0.230

Authors:

F.N.Musayev,F.Zarate-Perez,C.R.Escalante

Key ref:

F.N.Musayev et al. (2015). Structural Insights into the Assembly of the Adeno-associated Virus Type 2 Rep68 Protein on the Integration Site AAVS1. J Biol Chem, 290, 27487-27499. PubMed id: 26370092 DOI: 10.1074/jbc.M115.669960

Date:

08-May-15 Release date: 23-Sep-15

Protein chains

P03132  (REP68_AAV2S) -  Protein Rep68 from Adeno-associated virus 2 (isolate Srivastava/1982)

Seq: 536 a.a.

Struc: 185 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

DNA/RNA chains

G-C-G-C-T-C-G-C-T-C-G-C-T-C-G-C-T-G-G-G-C 21 bases

C-G-C-C-C-A-G-C-G-A-G-C-G-A-G-C-G-A-G-C-G 21 bases

Enzyme reactions

• Enzyme class: E.C.3.6.4.12 - Dna helicase.
• Reaction: ATP + H2O = ADP + phosphate + H⁺

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1074/jbc.M115.669960

J Biol Chem 290:27487-27499 (2015)

PubMed id: 26370092

Structural Insights into the Assembly of the Adeno-associated Virus Type 2 Rep68 Protein on the Integration Site AAVS1.

F.N.Musayev, F.Zarate-Perez, C.Bishop, J.W.Burgner, C.R.Escalante.

ABSTRACT

Adeno-associated virus (AAV) is the only eukaryotic virus with the property of establishing latency by integrating site-specifically into the human genome. The integration site known as AAVS1 is located in chromosome 19 and contains multiple GCTC repeats that are recognized by the AAV non-structural Rep proteins. These proteins are multifunctional, with an N-terminal origin-binding domain (OBD) and a helicase domain joined together by a short linker. As a first step to understand the process of site-specific integration, we proceeded to characterize the recognition and assembly of Rep68 onto the AAVS1 site. We first determined the x-ray structure of AAV-2 Rep68 OBD in complex with the AAVS1 DNA site. Specificity is achieved through the interaction of a glycine-rich loop that binds the major groove and an α-helix that interacts with a downstream minor groove on the same face of the DNA. Although the structure shows a complex with three OBD molecules bound to the AAVS1 site, we show by using analytical centrifugation and electron microscopy that the full-length Rep68 forms a heptameric complex. Moreover, we determined that a minimum of two direct repeats is required to form a stable complex and to melt DNA. Finally, we show that although the individual domains bind DNA poorly, complex assembly requires oligomerization and cooperation between its OBD, helicase, and the linker domains.