PDBsum entry 2iaj

Transferase

PDB id: 2iaj

Contents

Protein chains

551 a.a. *

415 a.a. *

Ligands

ATP

GOL ×2

Metals

_MN ×3

_NA

Waters ×308

* Residue conservation analysis

PDB id:

2iaj

Name:

Transferase

Title:

Crystal structure of k103n/y181c mutant HIV-1 reverse transcriptase (rt) in complex with atp

Structure:

Reverse transcriptase/ribonuclease h (p66 rt). Chain: a. Fragment: p66. Engineered: yes. Mutation: yes. Reverse transcriptase/ribonuclease h. P51 rt. Chain: b. Fragment: p51. Engineered: yes.

Source:

Human immunodeficiency virus type 1 bh10. Organism_taxid: 11678. Strain: bh10 isolate. Gene: pol. Expressed in: escherichia coli. Expression_system_taxid: 562. Expression_system_taxid: 562

Resolution:

2.50Å R-factor: 0.233 R-free: 0.284

Authors:

K.Das,E.Arnold

Key ref:

K.Das et al. (2007). Crystal Structures of Clinically Relevant Lys103Asn/Tyr181Cys Double Mutant HIV-1 Reverse Transcriptase in Complexes with ATP and Non-nucleoside Inhibitor HBY 097. J Mol Biol, 365, 77-89. PubMed id: 17056061 DOI: 10.1016/j.jmb.2006.08.097

Date:

08-Sep-06 Release date: 19-Dec-06

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: 551 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: 415 a.a.*

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

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

DOI no: 10.1016/j.jmb.2006.08.097

J Mol Biol 365:77-89 (2007)

PubMed id: 17056061

Crystal Structures of Clinically Relevant Lys103Asn/Tyr181Cys Double Mutant HIV-1 Reverse Transcriptase in Complexes with ATP and Non-nucleoside Inhibitor HBY 097.

K.Das, S.G.Sarafianos, A.D.Clark, P.L.Boyer, S.H.Hughes, E.Arnold.

ABSTRACT

Lys103Asn and Tyr181Cys are the two mutations frequently observed in patients exposed to various non-nucleoside reverse transcriptase inhibitor drugs (NNRTIs). Human immunodeficiency virus (HIV) strains containing both reverse transcriptase (RT) mutations are resistant to all of the approved NNRTI drugs. We have determined crystal structures of Lys103Asn/Tyr181Cys mutant HIV-1 RT with and without a bound non-nucleoside inhibitor (HBY 097, (S)-4-isopropoxycarbonyl-6-methoxy-3-(methylthio-methyl)-3,4-dihydroquinoxalin-2(1H)-thione) at 3.0 A and 2.5 A resolution, respectively. The structure of the double mutant RT/HBY 097 complex shows a rearrangement of the isopropoxycarbonyl group of HBY 097 compared to its binding with wild-type RT. HBY 097 makes a hydrogen bond with the thiol group of Cys181 that helps the drug retain potency against the Tyr181Cys mutation. The structure of the unliganded double mutant HIV-1 RT showed that Lys103Asn mutation facilitates coordination of a sodium ion with Lys101 O, Asn103 N and O(delta1), Tyr188 O(eta), and two water molecules. The formation of the binding pocket requires the removal of the sodium ion. Although the RT alone and the RT/HBY 097 complex were crystallized in the presence of ATP, only the RT has an ATP coordinated with two Mn(2+) at the polymerase active site. The metal coordination mimics a reaction intermediate state in which complete octahedral coordination was observed for both metal ions. Asp186 coordinates at an axial position whereas the carboxylates of Asp110 and Asp185 are in the planes of coordination of both metal ions. The structures provide evidence that NNRTIs restrict the flexibility of the YMDD loop and prevent the catalytic aspartate residues from adopting their metal-binding conformations.

Selected figure(s)

Figure 1.
Figure 1. Effects of the two mutations (Lys103Asn and Tyr181Cys) on the structure of unliganded HIV-1 RT. (a) A stereo view of the NNIBP region of the double mutant RT/ATP structure. The composite simulated annealing omit map (2|F[o]|–|F[c]|) electron density (cyan) contoured at 1.2σ defines the coordination of a Na ion at the NNIBP region; OW1 and OW2 are two water molecules. (b) The NNIBP region of the double mutant (Lys103Asn/Tyr181Cys) HIV-1 RT. The mutated amino acids have altered interactions with the surrounding amino acids. (c) The NNIBP region of the wild type unliganded HIV-1 RT structure.^13

Figure 2.
Figure 2. Binding mode of HBY 097 to the Lys103Asn/Tyr181Cys double mutant RT. (a) Stereo view of the (2|F[o]|–|F[c]|) electron density (contoured at 1.2σ) covering HBY 097 (cyan) and Cys181 (magenta). The dotted line represents the hydrogen bond between the thiol group of Cys181 and HBY 097. Electrostatic potential surface^62 showing the NNIBP region of (b) the double mutant RT/HBY 097 and (c) wild-type RT/HBY 097^13 structures.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2007, 365, 77-89) copyright 2007.

Figures were selected by an automated process.