PDBsum entry 1vsh

Endonuclease

PDB id: 1vsh

Contents

Protein chain

146 a.a. *

Ligands

EPE

Metals

_ZN ×4

Waters ×131

* Residue conservation analysis

PDB id:

1vsh

Name:

Endonuclease

Title:

Asv integrase core domain with zn(ii) cofactors

Structure:

Integrase. Chain: a. Fragment: catalytic core domain, residues 1 - 4, 52 - 209. Engineered: yes

Source:

Rous sarcoma virus (strain schmidt-ruppin). Organism_taxid: 11889. Strain: schmidt-ruppin. Expressed in: escherichia coli. Expression_system_taxid: 562. Other_details: original viral DNA clone: ju et al., J. Virol. 33:1026-1033 (1980), original expression clone terry et al., J. Virol. 62:2358-2365 (1988), expression clone for core: kulkosky et al., J. Virol. 206:448-456 (1995).

Biol. unit:

Dimer (from PDB file)

Resolution:

1.95Å R-factor: 0.173

Authors:

G.Bujacz,J.Alexandratos,A.Wlodawer

Key ref:

G.Bujacz et al. (1997). Binding of different divalent cations to the active site of avian sarcoma virus integrase and their effects on enzymatic activity. J Biol Chem, 272, 18161-18168. PubMed id: 9218451 DOI: 10.1074/jbc.272.29.18161

Date:

04-Mar-97 Release date: 15-May-97

Protein chain

O92956  (POL_RSVSB) -  Gag-Pol polyprotein from Rous sarcoma virus subgroup B (strain Schmidt-Ruppin)

Seq: 1603 a.a.

Struc: 146 a.a.*

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

Enzyme reactions

• Enzyme class 2: E.C.2.7.7.- - ?????
• Enzyme class 3: E.C.2.7.7.49 - RNA-directed Dna polymerase.
• Reaction: DNA(n) + a 2'-deoxyribonucleoside 5'-triphosphate = DNA(n+1) + diphosphate
• Enzyme class 4: E.C.2.7.7.7 - DNA-directed Dna polymerase.
• Reaction: DNA(n) + a 2'-deoxyribonucleoside 5'-triphosphate = DNA(n+1) + diphosphate
• Enzyme class 5: E.C.3.1.-.-
• Enzyme class 6: E.C.3.1.26.4 - ribonuclease H.
• Reaction: Endonucleolytic cleavage to 5'-phosphomonoester.
• Enzyme class 7: E.C.3.4.23.- - ?????

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

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

reference

DOI no: 10.1074/jbc.272.29.18161

J Biol Chem 272:18161-18168 (1997)

PubMed id: 9218451

Binding of different divalent cations to the active site of avian sarcoma virus integrase and their effects on enzymatic activity.

G.Bujacz, J.Alexandratos, A.Wlodawer, G.Merkel, M.Andrake, R.A.Katz, A.M.Skalka.

ABSTRACT

Retroviral integrases (INs) contain two known metal binding domains. The N-terminal domain includes a zinc finger motif and has been shown to bind Zn2+, whereas the central catalytic core domain includes a triad of acidic amino acids that bind Mn2+ or Mg2+, the metal cofactors required for enzymatic activity. The integration reaction occurs in two distinct steps; the first is a specific endonucleolytic cleavage step called "processing," and the second is a polynucleotide transfer or "joining" step. Our previous results showed that the metal preference for in vitro activity of avian sarcoma virus IN is Mn2+ > Mg2+ and that a single cation of either metal is coordinated by two of the three critical active site residues (Asp-64 and Asp-121) in crystals of the isolated catalytic domain. Here, we report that Ca2+, Zn2+, and Cd2+ can also bind in the active site of the catalytic domain. Furthermore, two zinc and cadmium cations are bound at the active site, with all three residues of the active site triad (Asp-64, Asp-121, and Glu-157) contributing to their coordination. These results are consistent with a two-metal mechanism for catalysis by retroviral integrases. We also show that Zn2+ can serve as a cofactor for the endonucleolytic reactions catalyzed by either the full-length protein, a derivative lacking the N-terminal domain, or the isolated catalytic domain of avian sarcoma virus IN. However, polynucleotidyl transferase activities are severely impaired or undetectable in the presence of Zn2+. Thus, although the processing and joining steps of integrase employ a similar mechanism and the same active site triad, they can be clearly distinguished by their metal preferences.

Selected figure(s)

Figure 2.
Fig. 2. A stereo view of the active site of ASV IN in the presence of divalent cations. Shown are the superimposed coordinates of the side chains of Asp-64 (D64), Asp-121 (D121), and Glu-157^ (E157) as well as the divalent cations. The complexes presented^ here are with Mn2+ (yellow), Zn2+ (green), Cd^2+ (red), and Ca^2+ (blue).

Figure 6.
Fig. 6. The environment of Mg2+ and Zn2+ divalent cations in the active site of polynucleotidyltransferases. The residues forming the D,D(35)E motif in the active site of ASV IN (green) are compared^ with the equivalent sites of HIV-1 RNase H (blue) as well as of^ the exonuclease domain of the Klenow fragment of DNA I polymerase^ (pink). One of the water molecules present in site I of ASV IN is found in a position occupied by carboxylate oxygens (O) in the other two structures. D, Asp-; E, Glu-.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (1997, 272, 18161-18168) copyright 1997.

Figures were selected by the author.