Part:BBa_K3515014

Alpha-Klotho Binding Protein with cysteine modification(s) to bind to a biosensor.

Alpha-Klotho selectively binds FGF23 (peptide hormone) in its active site. A fluorophore may be attached to the N- terminal domain as well to create a FRET signal. This makes it a distinguishable candidate for FGF23 monitoring using fluorescence resonance energy transfer (FRET). Coupling this protein with a supply of FGF23 bound to a second fluorophore can create a competition assay, where FGF23 and FGF23-fluorophore compete with one another for Alpha-Klotho. FGF23 detection is vital as FGF23 directly regulates renal function and is also a primary regulator of phosphate and vitamin D suppression. FGF23 is also a crucial biomarker used in clinical medicine for tracking the progression and status of Chronic Kidney Disease (CKD) patients. As such a biosensor for FGF23 tracking may be of great interest to patients and clinicians. This part includes Alpha-Klotho with a cysteine residue that will bind cysteine linker arms and be used for biosensor immobilization allowing the detection of FGF23. Disulfides bonds were not mutated and left in the protein construct.

Cysteine immobilization modifications of binding protein. Protein structures were obtained from the RCSB Protein Data Bank. All protein residues are shown in a blue cartoon preset with cysteine residues labelled in red cartoon, respectively, using PyMOL software.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 1626
  Illegal XhoI site found at 1165
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 160
  Illegal AgeI site found at 987
  Illegal AgeI site found at 1856
  
• 1000
  COMPATIBLE WITH RFC[1000]