HIV-1 reverse transcriptase

 

HIV-1 reverse transcriptase (RT) is a component of gag-pol polyprotein encoded by the human immunodeficiency virus-1 (HIV-1) genome. HIV-1 reverse transcriptase contains both DNA polymerase and ribonuclease H (RNase H) activities to convert the viral genomic RNA to double-stranded DNA, which is then integrated into the host genome and replicated along with the infected host cells. Understanding the HIV-1 reverse transcriptase catalytic mechanism is key for discovery of potent drugs and development of more effective, less toxic drugs and multi-drug combinations.

 

Reference Protein and Structure

Sequence
P03366 UniProt (2.7.7.-, 2.7.7.7, 2.7.7.49, 3.1.-.-, 3.1.13.2, 3.1.26.13, 3.4.23.16) IPR000477 (Sequence Homologues) (PDB Homologues)
Biological species
Human immunodeficiency virus type 1 BH10 (Virus) Uniprot
PDB
1rtd - STRUCTURE OF A CATALYTIC COMPLEX OF HIV-1 REVERSE TRANSCRIPTASE: IMPLICATIONS FOR NUCLEOSIDE ANALOG DRUG RESISTANCE (3.2 Å) PDBe PDBsum 1rtd
Catalytic CATH Domains
3.30.70.270 CATHdb (see all for 1rtd)
Cofactors
Magnesium(2+) (2)
Click To Show Structure

Enzyme Reaction (EC:2.7.7.49)

2'-deoxyribonucleoside 5'-triphosphate(4-)
CHEBI:61560ChEBI
+
DNA polyanion
CHEBI:83828ChEBI
→
diphosphate(3-)
CHEBI:33019ChEBI
+
DNA polyanion (n+1)
CHEBI:X00692X00692
Alternative enzyme names: DNA nucleotidyltransferase (RNA-directed), RNA revertase, RNA-dependent DNA polymerase, RNA-dependent deoxyribonucleate nucleotidyltransferase, RNA-instructed DNA polymerase, RT, Reverse transcriptase, Revertase,

Enzyme Mechanism

Introduction

HIV-1 reverse transcriptase mechanism consists of two main steps: deprotonation of the 3'-primer terminus by Asp185 followed by nucleophilic attack of the 3'OH towards the alpha-phosphate of the incoming deoxy-nucleotide (dNTP). The pyrophosphate leaving group is then protonated by Lys220.

Catalytic Residues Roles

UniProt PDB* (1rtd)
Asp784 Asp185A(E) Asp185 is responsible for the deprotonation of primer's 3'OH group metal ligand, proton acceptor
Asp709 Asp110A(E) Asp110 coordinates the Mg2+ metal ions metal ligand
Asp785 Asp186A(E) Asp186 coordinates the Mg2+ metal ions metal ligand
Val710 (main-C) Val111A(E) (main-C) Val111 coordinates the Mg2+ metal ions metal ligand
Lys819 Lys220A(E) Lys220 acts as a general acid for the protonation of pyrophosphate leaving group proton donor
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

proton transfer, coordination to a metal ion, overall product formed, overall reactant used, bimolecular nucleophilic substitution

References

  1. Rungrotmongkol T et al. (2014), Medchemcomm, 5, 593-. QM/MM simulations indicate that Asp185 is the likely catalytic base in the enzymatic reaction of HIV-1 reverse transcriptase. DOI:10.1039/c3md00319a.
  2. Castro C et al. (2009), Nat Struct Mol Biol, 16, 212-218. Nucleic acid polymerases use a general acid for nucleotidyl transfer. DOI:10.1038/nsmb.1540. PMID:19151724.
  3. Castro C et al. (2007), Proc Natl Acad Sci U S A, 104, 4267-4272. Two proton transfers in the transition state for nucleotidyl transfer catalyzed by RNA- and DNA-dependent RNA and DNA polymerases. DOI:10.1073/pnas.0608952104. PMID:17360513.
  4. Rungrotmongkol T et al. (2007), J Mol Graph Model, 26, 1-13. Active site dynamics and combined quantum mechanics/molecular mechanics (QM/MM) modelling of a HIV-1 reverse transcriptase/DNA/dTTP complex. DOI:10.1016/j.jmgm.2006.09.004. PMID:17046299.
  5. Huang H et al. (1998), Science, 282, 1669-1675. Structure of a covalently trapped catalytic complex of HIV-1 reverse transcriptase: implications for drug resistance. DOI:10.1126/science.282.5394.1669. PMID:9831551.

Step 1. Asp185 acts a general base to deprotonate the 3'-OH primer terminus. Active site residues serve as ligand to coordinate the metal ions Mg2+.

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Catalytic Residues Roles

Residue Roles
Asp185A(E) proton acceptor, metal ligand
Asp110A(E) metal ligand
Asp186A(E) metal ligand
Val111A(E) (main-C) metal ligand

Chemical Components

proton transfer, coordination to a metal ion

Step 2. Resulting hydroxide 3'-O performs nucleophilic attack towards the alpha-phosphate, leading to polymerization of nucleotides and hydrolysis of pyrophosphate. The pyrophosphate leaving group is protonated by Lys220.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp185A(E) metal ligand
Asp110A(E) metal ligand
Asp186A(E) metal ligand
Val111A(E) (main-C) metal ligand
Lys220A(E) proton donor

Chemical Components

coordination to a metal ion, overall product formed, overall reactant used, proton transfer, ingold: bimolecular nucleophilic substitution
Download: Image, Marvin File

Step 1. Asp185 acts a general base to deprotonate the 3'-OH primer terminus. Active site residues serve as ligand to coordinate the metal ions Mg2+.

Step 2. Resulting hydroxide 3'-O performs nucleophilic attack towards the alpha-phosphate, leading to polymerization of nucleotides and hydrolysis of pyrophosphate. The pyrophosphate leaving group is protonated by Lys220.

Products.

Contributors

Trung Nguyen, Antonio Ribeiro