MOESM1 of Recovery of bioactive protein from bacterial inclusion bodies using trifluoroethanol as solubilization agent

Additional file 1: Figure S1. Analysis of NATA fluorescence and acrylamide quenching of NATA. a) Fluorescence spectra of NATA incubated in different buffers. b) Stern–Volmer plots for acrylamide quenching of NATA in presence of different buffers

Proteinase K degradation profiles of hGH and asparaginase inclusion bodies isolated after 4 hours of induction.

<p>(<b>a</b>) Rate of degradation of IBs with time. (<b>b</b>) Rate of change in degradation rate of IBs with decrease in turbidities.</p

Transmission electron micrograph of purified hGH inclusion bodies.

<p>(<b>a</b>), (<b>b</b>), (<b>c</b>) and (<b>d</b>) are hGH IBs isolated after 1, 2, 3 and 4 hours of IPTG induction respectively. Bar represents 2 µm.</p

Amino acid sequence of hGH (Swiss-Prot, P01241).

<p>Bold and italics regions indicate presence of hydrophobic residues involves in making hydrophobic and amphipathic helices.</p

Purification, proteolytic digestion and solubilization profile of human growth hormone and asparaginase inclusion bodies.

<p>(a) Purification of inclusion bodies by sucrose density gradient ultracentrifugation, tube one : asparaginase IB, tube two: hGH inclusion bodies (b) SDS-PAGE analysis of purified hGH (21 kDa, lane 1) and asparaginase (37 kDa, lane 3) inclusion bodies. Lane 2 and 4, LMW marker: 97, 66, 45, 30, 20.1 and 14.4 kDa. (<b>c</b>) Kinetics of proteolytic digestion of inclusion body aggregates by Proteinase K. SDS-PAGE of solubilized inclusion body supernatants. (<b>d</b>) hGH IBs. Lane 1–9, supernatants of 0 to 8 molar urea. (<b>e</b>) Asparaginase IBs. Lane 1–9, supernatants of 0 to 8 molar urea; lane M, LMW marker (97, 66, 45, 30, 20.1 and 14.4 kDa).</p

Transmission electron micrograph of purified asparaginase inclusion bodies.

<p>(<b>a</b>), (<b>b</b>), (<b>c</b>) and (<b>d</b>) asparaginase IBs isolated after 1, 2, 3 and 4 hours of IPTG induction respectively. Bar represents 2 µm.</p

Relative activities of refolded asparaginase after solubilization of asparaginase IBs in different concentrations of urea.

<p>Relative activities of refolded asparaginase after solubilization of asparaginase IBs in different concentrations of urea.</p

Size distribution patterns of IBs isolated from <i>E. coli</i> cells harvested at different time points after induction.

<p>(<b>a</b>) Asparaginase IBs. (<b>b</b>) hGH IBs. Colored bars indicate the harvesting time.</p

Second derivative FTIR spectra of hGH and asparaginase IBs.

<p>(<b>a</b>) Second derivative spectrum of hGH IBs. (<b>b</b>) Second derivative spectrum of asparaginase IBs in amide band region. Broader peak in 1620–1640 cm-1 and 1680–1685 cm-1 range indicates large beta sheet content.</p

Solubilization profiles of inclusion bodies isolated from cell harvested at different time points after induction.

<p>(<b>a</b>) hGH IBs (<b>b</b>) Asparaginase IBs. Colored bars indicate the harvesting time.</p