1. Volume 1, Issue 2, Pages 77-89 (April 1996)
Mutational effects on inclusion body formation in the periplasmic expression of the immunoglobulin VL domain REI 
Winnie Chan, Larry R Helms, Ian Brooks, Grace Lee, Sarah Ngola, Dean McNulty, Beverly Maleeff, Preston Hensley, Ronald Wetzel 
Folding and Design 
Volume 1, Issue 2, Pages (April 1996)
DOI: /S (96)00017-X
Copyright © 1996 Elsevier Ltd Terms and Conditions

2. Figure 1
SDS polyacrylamide gels on subcellular fractionation of E. coli producing REI VL sequence variants. (a) Lanes 1 and 5, MW standards; lanes 2 and 6, REI WT, purified; lanes 3 and 7, tac vector minus REI gene; lanes 4 and 8, tac vector with WT REI gene; lanes 3 and 4, total cellular protein; lanes 7 and 8, osmotic shock fraction. (b) Native lysis supernatants and pellets showing distribution into IBs of various mutants. Lanes as marked. Loading of some pellet lanes is disproportionately high, compared with supernatant lanes, to improve quantitation.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
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3. Figure 2
Electron micrographs of REI-RGD4 VL expressed in the (a) cytoplasm and (b) periplasm of E. coli. i, inclusion body; arrowheads, periplasmic membrane; bars = μm.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
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4. Figure 3
Ribbon diagram of the structural model for the dimer of wild-type REI, from the X-ray coordinate set 1REI [23] in the Brookhaven database [46]. Chain A of the homodimer is in orange, chain B in green. Positions involving mutations described in this paper are further colored. Those at or near the dimer interface (residues 41–43 from RGD6; Q38, Q89, and CDR3 from mutants RGD4 and 12) are in shades of red, those distal to the interface (CDR1 portion altered in RGD34, R61) are in purple.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
Copyright © 1996 Elsevier Ltd Terms and Conditions

5. Figure 4
Equilibrium sedimentation data in the ultracentrifuge analysis of association behavior of wild-type REI VL. Data reported here were acquired at three rotor speeds at 25°C in a Beckman Optima XL-A analytical ultracentrifuge. The primary data (b) were obtained at 15, 20 and 25 000 rpm, respectively. Part (a) shows the distributions of residuals for the fit of the data to the model. The three panels in (a) are for 25, 20 and 15 000 rpm, respectively. The buffer was 20 mM sodium phosphate, 150 mM NaCl, pH 7.4. The initial protein concentration was ∼18 mM (in terms of monomer concentration, i.e. A280 ∼0.3). These data were fit best to a monomer/dimer model, equation 4. The value of K1,2 was determined to be 7.47 ± 0.96 × 10−6 M.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
Copyright © 1996 Elsevier Ltd Terms and Conditions

6. Figure 4
Equilibrium sedimentation data in the ultracentrifuge analysis of association behavior of wild-type REI VL. Data reported here were acquired at three rotor speeds at 25°C in a Beckman Optima XL-A analytical ultracentrifuge. The primary data (b) were obtained at 15, 20 and 25 000 rpm, respectively. Part (a) shows the distributions of residuals for the fit of the data to the model. The three panels in (a) are for 25, 20 and 15 000 rpm, respectively. The buffer was 20 mM sodium phosphate, 150 mM NaCl, pH 7.4. The initial protein concentration was ∼18 mM (in terms of monomer concentration, i.e. A280 ∼0.3). These data were fit best to a monomer/dimer model, equation 4. The value of K1,2 was determined to be 7.47 ± 0.96 × 10−6 M.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
Copyright © 1996 Elsevier Ltd Terms and Conditions

7. Figure 5
Sedimentation equilibrium data for REI-RGD4 in terms of equation 3b. Experimental conditions are the same as described in Figure 4. (a) Distribution of residuals. (b) Fit of the data to equation 3b. (c) Derived distribution of species as a function of the total monomer concentration as determined by the fit of the data to the model.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
Copyright © 1996 Elsevier Ltd Terms and Conditions

8. Figure 5
Sedimentation equilibrium data for REI-RGD4 in terms of equation 3b. Experimental conditions are the same as described in Figure 4. (a) Distribution of residuals. (b) Fit of the data to equation 3b. (c) Derived distribution of species as a function of the total monomer concentration as determined by the fit of the data to the model.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
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9. Figure 6
Sequence changes introduced to produce the RGD loop swap mutants described in this paper.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
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10. Figure 7
Correlation of thermodynamic stabilities of REI VL mutants with their tendencies to form IBs during secretion expression in E. coli at 37°C. Data from Table 2.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
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11. Figure 8
A model for off-pathway loss of soluble protein product in the secretion expression of REI VL in E. coli. See Discussion.
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
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12. Figure 9
Sequence of DNA in the tac expression vector including Shine–Dalgarno region, pelB signal peptide, and 5′ and 3′ termini of structural gene. The figure shows the DNA sequence of the 5′ untranslated region, which includes the reported Shine–Delgarno sequence (underlined), of the pectate lyase B gene and its signal sequence followed by the 5′ and 3′ junctions of the REI gene found in the tac expression vector. The coding sequence for REI used in this construction is identical to that described previously for the cytoplasmic expression of REI [24].
Folding and Design 1996 1, 77-89DOI: ( /S (96)00017-X)
Copyright © 1996 Elsevier Ltd Terms and Conditions