Rapid parallel expression in E. Coli and insect cells: analysis of five lef gene products of the Autographa californica multiple nuclear polyhedrosis virus (AcMNPV).
A number of strategies are emerging for the high throughput (HTP) expression of recombinant proteins to enable structural and functional study. Here we describe a workable HTP strategy based on parallel protein expression in E. coli and insect cells. Using this system we provide comparative expression data for five proteins derived from the Autographa californica polyhedrosis virus genome that vary in amino acid composition and in molecular weight. Although the proteins are part of a set of factors known to be required for viral late gene expression, the precise function of three of the five, late expression factors (lefs) 6, 7 and 10, is unknown. Rapid expression and characterisation has allowed the determination of their ability to bind DNA and shown a cellular location consistent with their properties. Our data point to the utility of a parallel expression strategy to rapidly obtain workable protein expression levels from many open reading frames (ORFs).
School of Animal and Microbial Sciences, The University of Reading, Reading RG6 6AJ, UK.
Xiaodong
X
Xu
Ian M
IM
Jones
D000818:Animals; D016367:Baculoviridae / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D017924:Nucleopolyhedroviruses / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera; D013997:Time Factors; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D011994:Recombinant Proteins; D014764:Viral Proteins; C087084:late expression factor 7, baculovirus