Functional identification of novel activities: activity-based selection of proteins from complete proteomes.
The identification of proteins with desired activities, especially from complex samples such as plasma and whole blood, is a continual challenge. We have developed a technology platform called Functional Identification of Novel Activities (FIoNA) to discover desired protein activities from complex biological samples. FIoNA uses immobilized libraries of combinatorial peptide ligands to purify and concentrate essentially all of the components of a complex mixture on ligands synthesized on individual beads. No depletion or prefractionation of the starting material is performed before it is incubated with the library, and no a priori knowledge of the active protein or of the ligand to which it binds is required. Instead, the protein-loaded beads are individually evaluated en masse in disease- relevant assays to identify proteins possessing a desired function. Beads associated with the activity are selected, and the ligand is sequenced and resynthesized in bulk on the original backbone for purification and characterization of the active component. Here we illustrate the use of FIoNA in a cell proliferation assay to detect a growth factor present in conditioned cell medium at nanogram/milliliter concentrations. We also have selected beads associated with hydrolysis of nerve agent analogs in assays performed in 100,000-well microtiter plates.
Sarkar
Jolly
J
Plasma Derivatives, Jerome Holland Laboratory, American Red Cross Biomedical Services, Rockville, MD 20855, USA.
Soukharev
Serguei
S
Lathrop
Julia Tait
JT
Hammond
David J
DJ
D000595:Amino Acid Sequence; D000818:Animals; D001798:Blood Proteins / Q000737:chemistry / Q000302:isolation & purification; D002417:Cattle; D002846:Chromatography, Affinity / Q000379:methods*; D020650:Combinatorial Chemistry Techniques / Q000379:methods*; D006801:Humans; D007202:Indicators and Reagents; D008024:Ligands; D051379:Mice; D019151:Peptide Library; D011485:Protein Binding; D011506:Proteins / Q000032:analysis* / Q000737:chemistry / Q000145:classification*; D020543:Proteome / Q000737:chemistry*; D013329:Structure-Activity Relationship
D001798:Blood Proteins; D007202:Indicators and Reagents; D008024:Ligands; D019151:Peptide Library; D011506:Proteins; D020543:Proteome