Table 2 A list of additives, their specific functions, and pertinent examples of proinsulin fusion protein-containing inclusion body washes

EDTA

A metal chelator to inactivate metalloproteases (Ritchie 2012)

Disrupts the lipopolysaccharide site of insertion into the bacterial outer membrane (Schnaitman 1971)

0.5 mM

Mackin (2014)

1 mM

Chen et al. (2016)

Hwang et al. (2016)

Nilsson et al. (1996)

Yuan et al. (2015)

2 mM

Leng et al. (2013)

Tikhonov et al. (2001)

Glycerol

NA

5%

Mackin (2014)

Lysozyme

Cleaves the backbone of peptidoglycan (Hwang et al. 2016)

0.02%

Hwang et al. (2016)

Kim et al. (2015)

Son et al. (2009)

NaCl

Solubilizes impurity through ionic interaction (Ritchie 2012)

0.05 M

Mackin (2014)

0.1 M

Leng et al. (2013)

0.2 M

Nilsson et al. (1996)

0.5 M

Mikiewicz et al. (2017)

Yuan et al. (2015)

Zieliński et al. (2019)

NA

Zimmerman and Stokell (2010)

Triton X-100

Removes membrane phospholipid and fragments (Singh et al. 2015; Palmer and Wingfield 2012)

Facilitates dissociation of debris and soluble proteins from inclusion bodies (Petrides et al. 1995)

0.66%

Petrides et al. (1995)

1%

Chen et al. (2016)

Hwang et al. (2016)

Kim et al. (2015)

Leng et al. (2013)

Mikiewicz et al. (2017)

Min et al. (2011)

Son et al. (2009)

Tikhonov et al. (2001)

Zieliński et al. (2019)

2%

Castellanos-Serra et al. (1996)

Cowley and Mackin (1997)

NA

Zimmerman and Stokell (2010)

Tween 20

Extracts the bacterial outer membrane components (Palmer and Wingfield 2012)

0.05%

Nilsson et al. (1996)

Zimmerman and Stokell (2010)

NA

Zimmerman and Stokell (2010)

Urea

A chaotropic agent to extract the bacterial outer membrane components (Palmer and Wingfield 2012; Petrides et al. 1995)

0.5 M

Hwang et al. (2016)

Kim et al. (2015)

2 M

Chen et al. (2016)

Cowley and Mackin (1997)

Kim et al. (2015)

Min et al. (2011)

Son et al. (2009)

Yuan et al. (2015)

3 M

Tikhonov et al. (2001)

4 M

Castellanos-Serra et al. (1996)