Archaeoglobus fulgidus RNase HII in DNA replication: enzymological functions and activity regulation via metal cofactors.

Abstract:

:RNA primer removal during DNA replication is dependent on ribonucleotide- and structure-specific RNase H and FEN-1 nuclease activities. A specific RNase H involved in this reaction has long been sought. RNase HII is the only open reading frame in Archaeoglobus fulgidus genome, while multiple RNases H exist in eukaryotic cells. Data presented here show that RNase HII from A. fulgidus (aRNase HII) specifically recognizes RNA-DNA junctions and generates products suited for the FEN-1 nuclease, indicating its role in DNA replication. Biochemical characterization of aRNase HII activity in the presence of various divalent metal ions reveals a broad metal tolerance with a preference for Mg(2+) and Mn(2+). Combined mutagenesis, biochemical competitions, and metal-dependent activity assays further clarify the functions of the identified amino acid residues in substrate binding or catalysis, respectively. These experiments also reveal that Asp129 form a second-metal binding site, and thus contribute to activity attenuation.

authors

Chai Q,Qiu J,Chapados BR,Shen B

doi

10.1006/bbrc.2001.5523

subject

Has Abstract

pub_date

2001-09-07 00:00:00

pages

1073-81

issue

5

eissn

0006-291X

issn

1090-2104

pii

S0006-291X(01)95523-8

journal_volume

286

pub_type

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