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Table 5 A summary of reported downstream purification schemes for recombinant human insulin/analogues and their precursors produced in E. coli

From: Downstream processing of recombinant human insulin and its analogues production from E. coli inclusion bodies

Fusion protein expressed in E. coli   Downstream purification schemes (purification steps arranged in sequence) References
rhPI IB recovery IMAC IEX RP    Astolfi et al. (2004)
rhPI IB recovery HIC (+ ⭮)        Bai et al. (2003)
rhPI IB recovery pH-PPT AEX pH-PPT RP RP Castellanos-Serra et al. (1996)
rhPI IB recovery AEX RP      Cowley and Mackin (1997)
rhPI IB recovery MMC Cryst CEX    GE app notes 28-9966-22 AA (2012), 29-0018-56 AB (2012), Heldin et al. (2014)
rhPI IB recovery pH-PPT pH-PPT     Kim et al. (2015)
rhPI   IEX RP SEC     Kroeff et al. (1989)
rhPI IB recovery AC RP      Mackin (1999)
rhPI IB recovery RP      Mackin and Choquette (2003)
rhPI IB recovery pH-PPT pH-PPT     Min et al. (2011)
rhPI IB recovery AC RP     Nilsson et al. (1996)
rhPI IB recovery CEX HIC IEX RP SEC Petrides et al. (1995)
rhPI IB recovery IMAC ZnCl2-PPT CEX    Redwan et al. (2007)
rhPI IB recovery IMAC        Yuan et al. (2015)
rhPI IB recovery SEC (+ ⭮)        Yuan et al. (2015)
rhPI IB recovery AEX ZnCl2-PPT AEX RP Zieliński et al. (2019)
rhPI, analogues   CEX       Coleman et al. (2019)
rhPI, analogues IB recovery AEX CEX      Thurow et al. (2010)
rhPI, analogues IB recovery AEX IMAC       Tikhonov et al. (2001)
rhPI, analogues IB recovery IMAC AEX       Tikhonov et al. (2001)
rhPI, analogues   MMC RP      Watson et al. (2018)
rhPI, analogues IB recovery AEX AEX RP   Mikiewicz et al. (2017)
PI analogues IB recovery IMAC RP     Zimmerman and Stokell (2010)
PI aspart IB recovery AEX       Chen et al. (2016)
PI glargine IB recovery pH-PPT CEX RP    Hwang et al. (2016)
Molecule
Inclusion body (IB) Precursor of human insulin/insulin analogue  Human insulin/insulin analogue
Chromatography
AC: affinity chromatography HIC: hydrophobic interaction chromatography MMC: mixed-mode chromatography
AEX: anion exchange chromatography IEX: ion-exchange chromatography RP: reversed-phase chromatography
CEX: cation exchange chromatography IMAC: immobilized metal ion affinity chromatography SEC: size exclusion chromatography
  1. Inclusion body (IB) (represented in bold italics format)
  2. Precursor of human insulin/insulin analogue (represented in italics format)
  3. Human insulin/insulin analogue (represented in underlined format)
  4. Cryst. crystallization, IB recovery protein recovery from inclusion bodies, pH-PPT pH precipitation, ZnCl2-PPT zinc chloride precipitation, rhPI Recombinant human proinsulin
  5. ⭮: Renaturation
  6. (+ ⭮): Simultaneous purification and renaturation
  7. ✁: Enzymatic conversion (e.g., Citraconylation & Trypsinization, Cleavage by trypsin and/or CPB)