Reverse transcriptase

Crystal Structure of unliganded HIV-1 Reverse transcriptase, 1hmv

Reverse transcriptase (RT) or RNA-dependent DNA polymerase transcribes single-stranded RNA into double-stranded DNA. HIV-1 RT is from the human immunodeficiency virus and is a heterodimer of P66 and P51. The images at the left and at the right correspond to one representative RT structure, i.e. crystal structure of HIV-1 Reverse transcriptase (1hmv). More details on HIV-1 RT in Phl p 2. P15 is its RNAse H domain. NNRTI are the non-nucleoside inhibitors of HIV-1 RT and NRTI are the nucleoide inhibitors. M-MLV is RT from the leukemia virus. Being the protein that gives their name to Retroviruses, Reverse Transcriptase is, in company of Protease and Integrase, the most important part of the protein system involved in the process of infection of viruses like HIV, MuLV and AMV, and has the unusual property of transcribing ssRNA into dsDNA going against the Central Dogma of Molecular Biology. Since its discovery in 1970, the study of its properties and mechanisms of action have been of high interest among the scientific community due to the unique properties that makes it an important medical target enzyme and important tool for genetic engineering applications like RT-PCR in the construction of cDNA libraries.

Reverse Transcriptase is one of the CBI Molecules being studied in the University of Massachusetts Amherst Chemistry-Biology Interface Program at UMass Amherst and on display at the Molecular Playground.

StructureStructure

spinqualitylabelsall models The hand-like two-enzymes-in-one protein that amazingly makes DNA from RNA Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

This hand-like protein that has an usual length of 1000 residues (560 in Chain A and 440 for B), the third of them involved in alpha helical and almost a quarter in beta sheets, showing α+β domains; has an usual weight of 66KDa whereas is around 51KDa, those monomers are derived from the same gene but p51 lacks the amino acids of one active site and has a different tertiary structure conformation compared with p66, for this reason is enzymatically inactive.

FunctionFunction

Template:STRUCTURE 3hvt As a RNA-dependent DNA Polymerase, is able to recognize the initial RNA, transcribe it to ssDNA, cleave the remaining RNA and then build up the dsDNA, to do this the protein has two active catalytic zones. Chain A has the that consist of two finger-like domains, one of them recognizes the initial nucleic acid by h-bonds interactions with phosphate groups of the side chains, then both domains make a conformational change closing the recognition hole to allow the second domain with the support a coordination system, begin the transcription process adding the specific DNA nucleotides; this change is allowed by a between the two previous domains that is used as a common pharmaceutical target site in order to prevent this change and by this way inhibit the activity, but this one is the only zone of the Chain A that has non-conserved aminoacids giving the virus more drug resistance. [1] As the same rate that the polymerization process occurs, the other active site known as the , cleaves the RNA releasing the ssDNA that comes again trough the Polymerase active site to become dsDNA (all this with a coordinative system, that allows non-specific recognition, just with phosphates). Finally Chain B despite of the similar aminoacid sequence with Chain A has no enzyme activity, possibly its function is to stabilize and communicate both active sites, this varying the length between them in order to synchronize both functions. This seems to be the most general idea of the mechanism of action of Reverse Transcriptase, however the process remains unclear and new approaches are being reported. [2]

One of the principal issues about this protein comparative with the usual DNA polymerase (besides to the similarity with the Klenow fragment), is the lack of a correction mechanism (usually made by DNA PolIII in the DNA Polymerase) that increase the quota of errors, producing more mutations and by this way giving more facultative and resistance abilities to the virus.

3D Structures of Reverse transcriptase3D Structures of Reverse transcriptase

Update December 2011

HIV-1 RT P66/P51HIV-1 RT P66/P51

3dlk, 1hmv - P66/P51
1mu2, 1jle, 1hpz, 1hqu, 1qe1, 1dlo - P66/P51 (mutant)
1har - P66 N-terminal
3qip - P66/P51+NNRTI nevirapine + RNase H inhibitor
3lp0, 3lp1, 3lp2, 1vrt - P66/P51+NNRTI nevirapine
2hnd, 2hny, 1s1u, 1s1x, 1lw0, 1lwc, 1lwe, 1lwf, 1jlb, 1jlf, 1fkp - P66/P51 (mutant)+NNRTI nevirapine
3m8p, 3mec - P66/P51+NNRTI etravirine
3med - P66/P51 (mutant)+NNRTI etravirine
3mee, 2zd1- P66/P51+NNRTI rilpivirine
3meg, 2ze2, 3bgr - P66/P51 (mutant)+NNRTI rilpivirine
1ikw, 1fk9 - P66/P51 +NNRTI efavirenz
1jkh, 1ikv, 1fko - P66/P51 (mutant)+NNRTI efavirenz
3dya, 3e01, 3drp - P66/P51+NNRTI pyrazole
3drr, 3drs - P66/P51 (mutant)+NNRTI pyrazole
3irx, 3is9, 3lak, 3lal, 3lam, 3lan, 3ffi, 3di6, 3dle, 3dlg, 2rki, 3c6t, 3c6u, 2rf2, 2vg5, 2vg6, 2vg7, 2opp, 1jlq, 1ep4, 1c0t, 1c0u, 1dtt, 1dtq, 1bqm, 1rt1, 1rt2, 1rev, 1rtj, 1vru, 1hnv, 1hni, 3t19 - P66/P51+NNRTI
1s1t, 1s1v, 1s1w, 1s6p, 1s6q, 1s9e, 1s9g, 1suq, 1sv5, 3dm2, 3dmj, 3dok, 3dol, 2opq, 2opr, 2ops, 2ic3, 2hnz, 1jlg, 1bqn, 1uwb, 1tvr, 3qo9, 3t1a - P66/P51 (mutant)+NNRTI
3kk1, 1c1b, 1rti - P66/P51+RTI
1jla - P66/P51 (mutant)+ NRTI
3ig1 - P66/P51+ inhibitor beta-thujaplicinol
3i0r, 3i0s, 3m8q, 3isn, 3ith, 1c1c, 3nbp - P66/P51+ pyrimidine derivative
1ikx - P66/P51 (mutant)+ pyrimidine derivative
1tv6 - P66/P51+ pyridine derivative
1jlc, 1iky, 1eet - P66/P51(mutant)+ pyridine derivative
2b5j, 2ban, 2be2, 1rth, 1hvt - P66/P51+ pyridone derivative
1lw2, 1rt3 - P66/P51 (mutant)+ NRTI
1tkt, 1tkx, 1tkz, 1tl1, 1tl3 - P66/P51+ quinoline derivative
2b6a - P66/P51+ THR-50
2i5j - P66/P51 (mutant)+hydrazone derivative
1rt4, 1rt5, 1rt6, 1rt7 - P66/P51+ carboxanilide derivative
1klm - P66/P51+piperazine derivative
2jle – P66+NNRTI
3kjv, 3kk3, 1r0a, 1n5y, 1n6q, 1hys - P66/P51+DNA
1j5o, 1rtd, 3klf - P66/P51 (mutant)+DNA
1hvu - P66/P51+RNA
1t03, 1t05 - P66/P51+DNA+Fab+NRTI
3jsm - P66/P51 (mutant)+DNA+NRTI
2hmi - P66/P51 (mutant)+DNA+Fab
2iaj - P66/P51 (mutant)+NRTI
3kk2 - P66/P51+DNA+NRTI
3jyt - P66/P51 (mutant)+DNA+NRTI

HIV-1 AZT-resistant RT P66/P51HIV-1 AZT-resistant RT P66/P51

3kli - P66/P51 (mutant)
3kle, 3klg, 3klh – P66/P51 (mutant) + DNA

HIV-1 RT P66HIV-1 RT P66

2jle – P66+NNRTI
1har - P66 N-terminal

HIV-1 RT P15HIV-1 RT P15

1rdh, 1hrh – P15
3lp3 – P15+inhibitor MK3
3k2p – P15+inhibitor beta-thujaplicinol
3hyf – P15+pyrimidine derivative
1o1w – P15+Mg - NMR

HIV-2 RTHIV-2 RT

1mu2 - RT (mutant)

Leukemia virus RTLeukemia virus RT

2hb5, 1mml - catalytic fragment
2r2r, 2r2t, 2fjv, 2fjw, 2fjx, 1ztw, 2fvs, 2fvr, 2fvq, 2fvp, 1n4l – catalytic fragment+DNA
2r2s, 2r2u - catalytic fragment+DNA+Co Bleomycin
1ztt - catalytic fragment+DNA+netropsin
1rw3, 1nnd – fragment (mutant)
1i6j, 1d0e, 1d1u, 1qaj, 1qai – N-terminal+DNA
3fsi - catalytic fragment+DNA+trypanocidal

RT from telomeraseRT from telomerase

2b2a - RT telomerase catalytic subunit (mutant) - Tetrahymena thermophila
2r4g - RT telomerase RNA-binding domain
3du6, 3du5 - TcRT telomerase catalytic subunit - Tribolium castaneum
3kyl - TcRT telomerase catalytic subunit + DNA

See AlsoSee Also

• Reverse Transcriptase at Wikipedia
• Molecule of the Month (09/2002) at RCSB Protein Data Bank
• List of Reverse Transcriptase articles at Proteopedia and at RCSB Protein Data Bank
• Model of Reverse Transcriptase as one of the CBI Molecules on the Molecular Playground
• See Transcription for additional Proteopedia articles on the subject.
• For additional information, see: Human Immunodeficiency Virus
• For additional information, see: Transcription and RNA Processing

ReferencesReferences

• [2] Consurf Server Data Base. Evolutionary conservation profile for Reverse Transcriptase PDB file 1JLB
• [3] Abbondanzieri, E.A. et al. Nature 453, 184-189 (2008) | doi:10.1038/nature06941