Patent ID: 8669074
Filing Date: 2014-03-11
Classification: C07K,C12N,C12Q,G01N

Abstract:
1. A chimeric phosphorylation indicator comprising a first fluorescent protein, a phosphoaminoacid binding domain (PAABD) comprising the forkhead-associated (FHA)2 sequence set forth in SEQ ID NO: 57, a protein kinase C (PKC)-phosphorylatable domain, and a second fluorescent protein, wherein the phosphorylatable domain is operably linked to the PAABD; wherein the first fluorescent protein and the second fluorescent protein are in operable linkage through the phosphorylatable domain and the PAABD; wherein the first fluorescent protein is different from the second fluorescent protein; wherein at least one of said first or said second fluorescent protein comprises a non-oligomerizing fluorescent protein; wherein the first fluorescent protein and the second fluorescent protein are selected from the group consisting of an Aequorea green fluorescent protein (GFP; SEQ ID NO: 2), a Discosoma red fluorescent protein (RFP; SEQ ID NO: 12), an enhanced GFP (EGFP; SEQ ID NO:4), an enhanced CFP (ECFP; SEQ ID NO:6), an ECFP(1-227) (amino acids 1 to 227 of SEQ ID NO:6), an EYFP-V68L/Q69K (SEQ ID NO: 10), an enhanced YFP (EYFP; SEQ ID NO:8), citrine (SEQ ID NO: 10 with Q69M), an I125R DsRed mutant (SEQ ID NO:12 with I125R) and a fluorescent protein (SEQ ID NO:2) having a mutation of an amino acid residue selected from A206K, L221K, F223R, or a combination thereof; wherein the first fluorescent protein and the second fluorescent protein exhibit a detectable resonance energy transfer when the first fluorescent protein is excited; and wherein the PKC-phosphorylatable domain and phosphoaminoacid binding domain emit weak or no fluorescent light.