Table 13 Reported optimized conditions for proinsulin (PI) refolding via dilution
Protein | Refolding yield (%) | Refolding duration | Refolding buffer | pH | Urea/ GdnHCl (M) | EDTA (mM) | Oxidants/reductants/redox couples | Other additives | Temp. (°C) | Protein concentration (mg/mL) | References |
|---|---|---|---|---|---|---|---|---|---|---|---|
rhPI | 99 | 18–19 h | 12 mM NaHCO3 | 10.8 | – | 0.2 | – | – | 7–8 | NA | Zieliński et al. (2019) |
rhPI | 85 | 12 h | NA | NA | – | – | BME (1.5 mol/mol of SO32−) | – | 8 | < 1 mg/mL | Petrides et al. (1995) |
rhPI | 43 | 1 h | 10 mM Tris/10 mM glycine | 7.5 | 43 mM GdnHCl | 1 | 1.8 molar ratio of GSH/GSSG | 10 µM Vectrase-P | 25 | 10 µM | Winter et al. (2002a) |
rhPI | NA | 16 h | 50 mM glycine/NaOH | 10.5 | – | – | BME (1.5 mol per mol of cysteine S-sulfonate) | – | 4 | 2 mg/mL | Cowley and Mackin (1997) |
rhPI | NA | Overnight | 50 mM glycine/NaOH | 10.5 | – | 1 | 1 mM GSH, 1 mM GSSG | – | 4 | 0.1 mg/mL | Mackin and Choquette (2003) |
rhPI | NA | 16 h | 10 mM glycine | 10.6 | 0.6 M urea | – | 0.3 mM BME | – | 15 | 1.5 mg/mL | Son et al. (2009) |
HGH-PI fusion protein | NA | 24 h | 1% glycine, 50 mM Tris–HCl, 100 mM NaCl | 9.3 | 3.2 M urea | – | 1 mM GSH, 1 mM GSSG | 5% glycerol, 0.2% PEG | 4 | NA mg/mL | Leng et al. (2013) |
His8–Arg–proinsulin | 60–70 | 10–30 min | 10 mM Tris, 10 mM glycine | 10–11 | 3 M GdnHCl or 4 M urea | 1 | 0.5 mM cysteine, 4.5 mM cystine | – | 15 | 0.5 mg/mL | Winter et al. (2002b) |
Hybrid insulin analogue precursor | NA | 18 h | 2 mM NaHCO3 | 11.2 | – | 0.2 | – | – | NA | NA | Mikiewicz et al. (2017) |
Interleukin-2-proinsulin S-hexasulfonate | 80 | 6 h | 50 mM glycine | 10.5 | – | 1 | BME (1.5 eq. per eq. of S-sulfonate) | – | 4 | 0.1 mg/mL | Castellanos-Serra et al. (1996) |
Novel PI | NA | ≥ 48 h | 5 mM CAPS | 10.5 | – | – | Equal amount of GSSG to the initial amount of reducing agent | – | 2–10 | 0.5 mg/mL | Zimmerman and Stokell 2010) |
PI aspart | 80 | Overnight | 20 mM glycine–NaOH | 10 | 2 M urea | – | 10 µM oxidant of CASeSeCA | – | 4 | 0.3 mg/mL | Chen et al. (2016) |
PI aspart | 80 | Overnight | 20 mM glycine–NaOH | 10 | – | – | 5 µM of an oxidative agent of selenocystamine | 0.5 M arginine | 4 | 0.5 mg/mL | Chen et al. (2016) |
Prepeptide fusion glargine | 82.1 | 48 h | 50 mM glycine | 10.6 | 0.6 M urea | – | 0.1 mM BME | – | 4 | 0.5 mg/mL | Hwang et al. (2016) |
Pure sulfonated PI | NA | 18–24 h | 10 mM glycine | 10.0 | – | – | 0.5 mM cystine, 0.5 mM BME | – | 4–8 | 0.5 mg/mL | Astolfi et al. (2004) |
S-sulfonated fusion preproinsulin | 85 | 17 h | 50 mM glycine | 10.6 | 0.3 M urea | – | BME (0.75 equiv. of –SH per –SSO3−) | – | 12 | 0.5 mg/mL | Min et al. (2011) |
S-sulfonated PI | NA | 16 h | 50 mM glycine/NaOH | 10.5 | 1 M urea | 1 mM BME, 0.15 mM cystine | – | 4 | 2 mg/mL | Redwan et al. (2007) | |
S-sulfonated preproinsulin | 74.2 | 16 h | 50 mM glycine | 10.6 | 0.6 M urea | – | 0.3 mM BME | – | 15 | 1.5 mg/mL | Kim et al. (2015) |
ZZ-R-PI | NA | 20 h | 0.1 M glycine–NaOH | 10.5 | – | – | BME (18 mol/mol fusion protein) | – | 4 | 0.8 mg/mL | Nilsson et al. (1996) |