Patent ID: 8735559
Filing Date: 2014-05-27
Classification: A01K,A61K,C12N,C12Q,C12Y

Abstract:
1. A polynucleotide encoding a biosensor polypeptide comprising a modified circularly-permuted thermostable luciferase of a parental thermostable luciferase that is modified by introduction of one or more amino acid substitutions relative to the amino acid sequence of the parental thermostable luciferase wherein the circularly-permuted thermostable luciferase has a C-terminal portion of the thermostable luciferase joined to an N-terminal portion of the thermostable luciferase by a peptide linker comprising a sensor region capable of interacting with a target molecule in a cell, and wherein the modified circularly-permuted thermostable luciferase has an enhanced response after interaction of the biosensor with the target molecule relative to at least one of: i) the parental circularly-permuted thermostable luciferase in the presence of the target molecule, or ii) the modified circularly-permuted thermostable luciferase in the absence of the target molecule, wherein the modified circularly-permuted thermostable luciferase comprises a substitution of an amino acid in at least one position that corresponds to positions 193, 471, or 507 of SEQ ID NO:2, wherein the substitutions are S193P, I471T, and/or T507I, or a combination thereof, and wherein the encoded amino acid sequence of the parental thermostable luciferase has at least 90% identity to the amino acid sequence of SEQ ID NO:4.