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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)