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Table 1 The recombinant proteins of this study. The immobilised ligand used for affinity chromatography purification and the elution conditions are shown. Protein eluted using acetic acid was immediately dialyzed against 50 mM NaPO4 pH 7.0 + 100 mM NaCl.

From: Avidin related protein 2 shows unique structural and functional features among the avidin protein family

Protein Affinity chromatography ligand Elution conditions Expression system Production yield (mg/l)a
AVR2 D-biotin 0.5 M acetic acid BEVSb 0.8
AVR2-b D-biotin 0.5 M acetic acid E. coli c 4.9
AVR2(I109K) D-biotin 0.5 M acetic acid BEVS 9.0
AVR4d 2-iminobiotin 50 mM Na-Ac + 100 mM NaCl BEVS 6.1
AVR4-bd 2-iminobiotin 50 mM Na-Ac + 100 mM NaCl E. coli 21.9
AVR4(K109I)d 2-iminobiotin 50 mM Na-Ac + 100 mM NaCl BEVS 16.2
AVR4(K109I)-bd 2-iminobiotin 50 mM Na-Ac + 100 mM NaCl E. coli 6.7
AVR6-be D-biotin 2 M acetic acid E. coli 8.3
AVD(K111I) 2-iminobiotin 50 mM Na-Ac + 100 mM NaCl BEVS 11.5
AVD(K111I)-b 2-iminobiotin 50 mM Na-Ac + 100 mM NaCl E. coli 0.7
  1. aAfter affinity chromatography per one liter of culture medium. The yields are calculated based on only a few protein purifications.
  2. bProduced using a baculovirus expression system in insect cells as described previously [40].
  3. cThe bacterial signal peptide from the protein OmpA was utilised in order to produce the protein in an active form in E. coli as described in [37].
  4. dAVR4 is identical to AVR5. The protein carries the mutation C122S, which prevents oligomerisation via intermolecular disulphide bridges [36].
  5. eCarries mutation C58S, which prevents oligomerisation via intermolecular disulphide bridges (this study).