PDBsum entry 1r0a

Transferase/immune system/DNA

PDB id: 1r0a

Contents

Protein chains

558 a.a. *

429 a.a. *

211 a.a. *

225 a.a. *

DNA/RNA

Ligands

GOL ×2

GLC

Metals

_MG

Waters ×28

* Residue conservation analysis

PDB id:

1r0a

Name:

Transferase/immune system/DNA

Title:

Crystal structure of HIV-1 reverse transcriptase covalently tethered to DNA template-primer solved to 2.8 angstroms

Structure:

5'-d( A Tp Gp Cp Ap Tp Cp Gp Gp Cp Gp Cp Tp Cp Gp Ap Ap Cp Ap Gp Gp Gp Ap Cp Gp Gp T)-3'. Chain: t. Engineered: yes. Other_details: oligonucleotide DNA template. 5'-d( C Cp Gp Tp Cp Cp Cp Tp Gp Tp Tp Cp Gp Ap Gp Cp Gp Cp Cp Gp (2Da))-3'. Chain: p. Engineered: yes.

Source:

Synthetic: yes. Human immunodeficiency virus 1. Organism_taxid: 11676. Gene: pol. Expressed in: escherichia coli. Expression_system_taxid: 562. Mus musculus. House mouse. Organism_taxid: 10090.

Biol. unit:

Hexamer (from PQS)

Resolution:

2.80Å R-factor: 0.239 R-free: 0.272

Authors:

S.Tuske,J.Ding,E.Arnold

Key ref:

E.N.Peletskaya et al. (2004). Nonnucleoside inhibitor binding affects the interactions of the fingers subdomain of human immunodeficiency virus type 1 reverse transcriptase with DNA. J Virol, 78, 3387-3397. PubMed id: 15016861

Date:

19-Sep-03 Release date: 03-Aug-04

Protein chain

P03366  (POL_HV1B1) -  Gag-Pol polyprotein from Human immunodeficiency virus type 1 group M subtype B (isolate BH10)

Seq: 1447 a.a.

Struc: 558 a.a.*

Protein chain

P03366  (POL_HV1B1) -  Gag-Pol polyprotein from Human immunodeficiency virus type 1 group M subtype B (isolate BH10)

Seq: 1447 a.a.

Struc: 429 a.a.*

Protein chain

No UniProt id for this chain

Struc: 211 a.a.

Protein chain

P01868  (IGHG1_MOUSE) -  Ig gamma-1 chain C region secreted form from Mus musculus

Seq: 324 a.a.

Struc: 225 a.a.*

* PDB and UniProt seqs differ at 6 residue positions (black crosses)

DNA/RNA chains

T-G-C-A-T-C-G-G-C-G-C-T-C-G-A-A-C-A-G-G-G-A-C-G 24 bases

C-G-T-C-C-C-T-G-T-T-C-G-A-G-C-G-C-C-G-2DA 20 bases

Enzyme reactions

• Enzyme class 1: Chains A, B: E.C.2.7.7.- - ?????
• Enzyme class 2: Chains A, B: E.C.2.7.7.49 - RNA-directed Dna polymerase.
• Reaction: DNA(n) + a 2'-deoxyribonucleoside 5'-triphosphate = DNA(n+1) + diphosphate
• Enzyme class 3: Chains A, B: E.C.2.7.7.7 - DNA-directed Dna polymerase.
• Reaction: DNA(n) + a 2'-deoxyribonucleoside 5'-triphosphate = DNA(n+1) + diphosphate
• Enzyme class 4: Chains A, B: E.C.3.1.-.-
• Enzyme class 5: Chains A, B: E.C.3.1.13.2 - exoribonuclease H.
• Reaction: Exonucleolytic cleavage to 5'-phosphomonoester oligonucleotides in both 5'- to 3'- and 3'- to 5'-directions.
• Enzyme class 6: Chains A, B: E.C.3.1.26.13 - retroviral ribonuclease H.
• Enzyme class 7: Chains A, B: E.C.3.4.23.16 - HIV-1 retropepsin.
• Reaction: Specific for a P1 residue that is hydrophobic, and P1' variable, but often Pro.

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

J Virol 78:3387-3397 (2004)

PubMed id: 15016861

Nonnucleoside inhibitor binding affects the interactions of the fingers subdomain of human immunodeficiency virus type 1 reverse transcriptase with DNA.

E.N.Peletskaya, A.A.Kogon, S.Tuske, E.Arnold, S.H.Hughes.

ABSTRACT

Site-directed photoaffinity cross-linking experiments were performed by using human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) mutants with unique cysteine residues at several positions (i.e., positions 65, 67, 70, and 74) in the fingers subdomain of the p66 subunit. Since neither the introduction of the unique cysteine residues into the fingers nor the modification of the SH groups of these residues with photoaffinity cross-linking reagents caused a significant decrease in the enzymatic activities of RT, we were able to use this system to measure distances between specific positions in the fingers domain of RT and double-stranded DNA. HIV-1 RT is quite flexible. There are conformational changes associated with binding of the normal substrates and nonnucleoside RT inhibitors (NNRTIs). Cross-linking was used to monitor intramolecular movements associated with binding of an NNRTI either in the presence or in the absence of an incoming deoxynucleoside triphosphate (dNTP). Binding an incoming dNTP at the polymerase active site decreased the efficiency of cross-linking but caused only modest changes in the preferred positions of cross-linking. This finding suggests that the fingers of p66 are closer to an extended template in the "open" configuration of the enzyme with the fingers away from the active site than in the closed configuration with the fingers in direct contact with the incoming dNTP. NNRTI binding caused increased cross-linking in experiments with diazirine reagents (especially with a diazirine reagent with a longer linker) and a moderate shift in the preferred sites of interaction with the template. Cross-linking occurred closer to the polymerase active site for RTs modified at positions 70 and 74. The effects of NNRTI binding were more pronounced in the absence of a bound dNTP; pretreatment of HIV-1 RT with an NNRTI reduced the effect of dNTP binding. These observations can be explained if the binding of NNRTI causes a decrease in the flexibility in the fingers subdomain of RT-NNRTI complex and a decrease in the distance from the fingers to the template extension.