Rapid isolation of novel FK506 binding proteins from multiple organisms using gDNA and cDNA T7 phage display.
Reverse chemical proteomics using T7 phage display is a powerful technique for identifying cellular receptors of biologically active small molecules. However, to date this method has generally been limited to cDNA libraries constructed from mRNA isolated from eukaryotes. In this paper, we describe the construction of the first prokaryotic T7 phage display libraries from randomly digested Pseudomonas stutzeri and Vibrio fischeri gDNA, as well as a plant cDNA library from Arabidopsis thaliana. We also describe the use of T7 phage display to identify novel proteins from environmental DNA samples using biotinylated FK506 as a model affinity probe.
Piggott
Andrew M
AM
Department of Chemistry and Biomolecular Sciences, Macquarie University, Sydney, NSW 2109, Australia.
Kriegel
Alison M
AM
Willows
Robert D
RD
Karuso
Peter
P
D048908:Aliivibrio fischeri / Q000235:genetics; D017360:Arabidopsis / Q000235:genetics; D017123:Bacteriophage T7; D053938:DNA Fragmentation; D004269:DNA, Bacterial; D018744:DNA, Plant; D015723:Gene Library; D007166:Immunosuppressive Agents; D015336:Molecular Probe Techniques; D015335:Molecular Probes; D019151:Peptide Library; D044223:Pseudomonas stutzeri / Q000235:genetics; D016559:Tacrolimus / Q000378:metabolism; D022021:Tacrolimus Binding Proteins / Q000302:isolation & purification*
D004269:DNA, Bacterial; D018744:DNA, Plant; D007166:Immunosuppressive Agents; D015335:Molecular Probes; D019151:Peptide Library; D022021:Tacrolimus Binding Proteins; D016559:Tacrolimus