Phage display of intact domains at high copy number: A system based on SOC, the small outer capsid protein of bacteriophage T4

Peptides fused to the coat proteins of filamentous phages have found widespread applications in antigen display, the construction of antibody libraries, and biopanning. However, such systems are limited in terms of the size and number of the peptides that may be incorporated without compromising the...

Full description

Saved in:

Bibliographic Details
Published in:	Protein science Vol. 5; no. 9; pp. 1833 - 1843
Main Authors:	Ren, Z.J., Black, L.W., Lewis, G.K., Wingfield, P.T., Locke, E.G., Steven, A.C.
Format:	Journal Article
Language:	English
Published:	Bristol Cold Spring Harbor Laboratory Press 01-09-1996
Subjects:	AIDS/HIV Animals Antigens, Viral - immunology bacteriophage T4 Bacteriophage T4 - chemistry Bacteriophage T4 - immunology Bacteriophage T4 - metabolism Capsid - chemistry Capsid - genetics Capsid - immunology Capsid - metabolism capsid assembly Capsid Proteins Escherichia coli - genetics Gene Expression Genetic Vectors HIV Envelope Protein gp120 - genetics HIV Envelope Protein gp120 - immunology human immunodeficiency virus antigens Mice Mice, Inbred BALB C phage display protein engineering Recombinant Fusion Proteins - immunology
Online Access:	Get full text
Tags:	Add Tag No Tags, Be the first to tag this record!

Description
Summary:	Peptides fused to the coat proteins of filamentous phages have found widespread applications in antigen display, the construction of antibody libraries, and biopanning. However, such systems are limited in terms of the size and number of the peptides that may be incorporated without compromising the fusion proteins' capacity to self‐assemble. We describe here a system in which the molecules to be displayed are bound to pre‐assembled polymers. The polymers are T4 capsids and polyheads (tubular capsid variants) and the display molecules are derivatives of the dispensable capsid protein SOC. In one implementation, SOC and its fusion derivatives are expressed at high levels in Escherichia coli, purified in high yield, and then bound in vitro to separately isolated polyheads. In the other, a positive selection vector forces integration of the modified soc gene into a soc‐deleted T4 genome, leading to in vivo binding of the display protein to progeny virions. The system is demonstrated as applied to C‐terminal fusions to SOC of (1) a tetrapeptide; (2) the 43‐residue V3 loop domain of gpl20, the human immunodeficiency virus type‐1 (HIV‐1) envelope glycoprotein; and (3) poliovirus VP1 capsid protein (312 residues). SOC‐V3 displaying phage were highly antigenic in mice and produced antibodies reactive with native gp120. That the fusion protein binds correctly to the surface lattice was attested in averaged electron micrographs of polyheads. The SOC display system is capable of presenting up to ∼ 103 copies per capsid and >104 copies per polyhead of V3‐sized domains. Phage displaying SOC‐VP1 were isolated from a 1:106 mixture by two cycles of a simple biopanning procedure, indicating that proteins of at least 35 kDa may be accommodated.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0961-8368 1469-896X
DOI:	10.1002/pro.5560050909