Structure of Escherichia coli O157:H7 bacteriophage CBA120 tailspike protein 4 baseplate anchor and tailspike assembly domains (TSP4-N)

Kinlin L Chao,Osnat Herzberg,Julia Greenfield,Adit B Alreja,Sara B Linden,Daniel C Nelson,Xiaoran Shang

doi:10.1038/s41598-022-06073-2

Abstract

Four tailspike proteins (TSP1-4) of Escherichia coli O157:H7 bacteriophage CBA120 enable infection of multiple hosts. They form a branched complex that attaches to the tail baseplate. Each TSP recognizes a different lipopolysaccharide on the membrane of a different bacterial host. The 335 N-terminal residues of TSP4 promote the assembly of the TSP complex and anchor it to the tail baseplate. The crystal structure of TSP4-N335 reveals a trimeric protein comprising four domains. The baseplate anchor domain (AD) contains an intertwined triple-stranded β-helix. The ensuing XD1, XD2 and XD3 β-sheet containing domains mediate the binding of TSP1-3 to TSP4. Each of the XD domains adopts the same fold as the respective XD domains of bacteriophage T4 gp10 baseplate protein, known to engage in protein–protein interactions via its XD2 and XD3 domains. The structural similarity suggests that XD2 and XD3 of TSP4 also function in protein–protein interactions. Analytical ultracentrifugation analyses of TSP4-N335 and of domain deletion proteins showed how TSP4-N335 promotes the formation of the TSP quaternary complex. TSP1 and TSP2 bind directly to TSP4 whereas TSP3 binding requires a pre-formed TSP4-N335:TSP2 complex. A 3-dimensional model of the bacteriophage CBA120 TSP complex has been developed based on the structural and ultracentrifuge information.

Highlights

Bacteriophage vB_EcoM_CBA120, discovered by Kutter and colleagues[1], is a member of the recently defined Kuttervirus genus of contractile tailed phages belonging to the Ackermannviridae family
The SeMet TSP4-N335 containing three constitutive methionines and three additional engineered methionines designed to increase the anomalous signal (Leu12Met, Ile31Met and Leu145Met), were all twinned when grown in KNa-tartrate solution
The three resolved SeMet-TSP4-N335 domains were used as search models for molecular replacement with diffraction data from

Summary

Introduction

Bacteriophage (phage) vB_EcoM_CBA120, discovered by Kutter and colleagues[1], is a member of the recently defined Kuttervirus genus of contractile tailed phages belonging to the Ackermannviridae family. The gp[66] TSP from the E. coli bacteriophage G7C contains a N-terminal region that attaches to the baseplate, followed by XD2 and XD3 domains Based on this homology between CBA120 TSP4 and gp[66] TSP, Plattner and colleagues inferred that the eighty N-terminal amino acid residues of TSP4 bind to the tail baseplate of phage CBA120 even though the N-terminal regions of gp[66] and TSP4 lack sequence homology. We refer to this baseplate anchor domain as AD (Fig. 1). The emerging model serves as a paradigm for the branched assemblies of TSPs from other Kuttervirus genus members

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Conclusion