title: Epitope discovery for a synthetic polymer nanoparticle: a new strategy for developing a peptide tag.
abstract: We describe a novel epitope discovery strategy for creating an affinity agent/peptide tag pair. A synthetic polymer nanoparticle (NP) was used as the "bait" to catch an affinity peptide tag. Biotinylated peptide tag candidates of varied sequence and length were attached to an avidin platform and screened for affinity against the polymer NP. NP affinity for the avidin/peptide tag complexes was used to provide insight into factors that contribute NP/tag binding. The identified epitope sequence with an optimized length (tMel-tag) was fused to two recombinant proteins. The tagged proteins exhibited higher NP affinity than proteins without tags. The results establish that a fusion peptide tag consisting of optimized 15 amino acid residues can provide strong affinity to an abiotic polymer NP. The affinity and selectivity of NP/tMel-tag interactions were exploited for protein purification in conjunction with immobilized metal ion/His6-tag interactions to prepare highly purified recombinant proteins. This strategy makes available inexpensive, abiotic synthetic polymers as affinity agents for peptide tags and provides alternatives for important applications where more costly affinity agents are used.
authors.affiliation: Department of Chemistry, University of California , Irvine, California 92697, United States.
authors.forename: Keiichi
authors.initials: K
authors.lastname: Yoshimatsu
authors.forename: Tomohiko
authors.initials: T
authors.lastname: Yamazaki
authors.forename: Yu
authors.initials: Y
authors.lastname: Hoshino
authors.forename: Paul E
authors.initials: PE
authors.lastname: Rose
authors.forename: Linda F
authors.initials: LF
authors.lastname: Epstein
authors.forename: Les P
authors.initials: LP
authors.lastname: Miranda
authors.forename: Philip
authors.initials: P
authors.lastname: Tagari
authors.forename: John M
authors.initials: JM
authors.lastname: Beierle
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Yonamine
authors.forename: Kenneth J
authors.initials: KJ
authors.lastname: Shea
mesh_terms: D001360:Avidin / Q000737:chemistry; D000939:Epitopes / Q000737:chemistry*; D015394:Molecular Structure; D053758:Nanoparticles / Q000737:chemistry*; D010316:Particle Size; D010455:Peptides / Q000737:chemistry*; D011108:Polymers / Q000138:chemical synthesis* / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D013499:Surface Properties
chemical_list: D000939:Epitopes; D010455:Peptides; D011108:Polymers; D011994:Recombinant Proteins; D001360:Avidin