Crystal Structure of PriB, a Primosomal DNA Replication Protein of Escherichia coli

Jyung-Hurng Liu,Tsai-Wang Chang,Cheng-Yang Huang,Sue-Une Chen,Huey-Nan Wu,Ming-Chung Chang,Chwan-Deng Hsiao

doi:10.1074/jbc.m406773200

Abstract

PriB is one of the Escherichia coli varphiX-type primosome proteins that are required for assembly of the primosome, a mobile multi-enzyme complex responsible for the initiation of DNA replication. Here we report the crystal structure of the E. coli PriB at 2.1 A resolution by multi-wavelength anomalous diffraction using a mercury derivative. The polypeptide chain of PriB is structurally similar to that of single-stranded DNA-binding protein (SSB). However, the biological unit of PriB is a dimer, not a homotetramer like SSB. Electrophoretic mobility shift assays demonstrated that PriB binds single-stranded DNA and single-stranded RNA with comparable affinity. We also show that PriB binds single-stranded DNA with certain base preferences. Based on the PriB structural information and biochemical studies, we propose that the potential tetramer formation surface and several other regions of PriB may participate in protein-protein interaction during DNA replication. These findings may illuminate the role of PriB in varphiX-type primosome assembly.

Highlights

A bacterial primosome is a multi-enzyme complex that participates in DNA replication
Based on the PriB structural information and biochemical studies, we propose that the potential tetramer formation surface and several other regions of PriB may participate in protein-protein interaction during DNA replication
The assembly of a primosome on the ␾X174 viral DNA, as described in the following sentences, is an ordered process [16]. (i) PriA recognizes and binds to the pas. (ii) PriB joins PriA to form a PriA-PriB-pas DNA complex. (iii) DnaT joins this complex to form a triprotein complex on the pas DNA; and (iv) DnaB is transferred from a DnaB-DnaC complex to the PriA-PriBDnaT-pas DNA complex to form a preprimosome assembly that consists of PriA, PriB, DnaT, and DnaB on the DNA

Summary

Introduction

A bacterial primosome is a multi-enzyme complex that participates in DNA replication. The polypeptide chain of PriB is structurally similar to that of single-stranded DNA-binding protein (SSB). Based on the PriB structural information and biochemical studies, we propose that the potential tetramer formation surface and several other regions of PriB may participate in protein-protein interaction during DNA replication.

Results

Conclusion