Chaperones activate hepadnavirus reverse transcriptase by transiently exposing a C-proximal region in the terminal protein domain that contributes to epsilon RNA binding.

Abstract:

:All hepatitis B viruses replicate by protein-primed reverse transcription, employing a specialized reverse transcriptase, P protein, that carries a unique terminal protein (TP) domain. To initiate reverse transcription, P protein must bind to a stem-loop, epsilon, on the pregenomic RNA template. TP then provides a Y residue for covalent attachment of the first nucleotide of an epsilon-templated DNA oligonucleotide (priming reaction) that serves to initiate full-length minus-strand DNA synthesis. epsilon binding requires the chaperone-dependent conversion of inactive P protein into an activated, metastable form designated P*. However, how P* differs structurally from P protein is not known. Here we used an in vitro reconstitution system for active duck hepatitis B virus P combined with limited proteolysis, site-specific antibodies, and defined P mutants to structurally compare nonactivated versus chaperone-activated versus primed P protein. The data show that Hsp70 action, under conditions identical to those required for functional activation, transiently exposes the C proximal TP region which is, probably directly, involved in epsilon RNA binding. Notably, after priming and epsilon RNA removal, a very similar new conformation appears stable without further chaperone activity; hence, the activation of P protein is triggered by energy-consuming chaperone action but may be completed by template RNA binding.

journal_name

J Virol

journal_title

Journal of virology

authors

Stahl M,Beck J,Nassal M

doi

10.1128/JVI.01196-07

subject

Has Abstract

pub_date

2007-12-01 00:00:00

pages

13354-64

issue

24

eissn

0022-538X

issn

1098-5514

pii

JVI.01196-07

journal_volume

81

pub_type

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