Stepwise purification of L-asparaginase from <i>Bacillus licheniformis.</i>

<p>Stepwise purification of L-asparaginase from <i>Bacillus licheniformis.</i></p

Anti-cancerous effect of enzyme on different human cancer cell lines <i>viz</i>. Jurkat clone E6-1, MCF-7 and K-562.

<p>Anti-cancerous effect of enzyme on different human cancer cell lines <i>viz</i>. Jurkat clone E6-1, MCF-7 and K-562.</p

Fluorescence spectroscopy showing the emission spectrum in the range of 300–400

<p> <b>nm (excitation wavelength: 292 nm) and unfolding transitions of L-asparaginase at 0 </b><b>M, 3 </b><b>M and 6 </b><b>M guanidine HCl.</b></p

Effect of chemical parameters on purified L-asparaginase a) Effect of metal ions (100 mM) on enzyme activity; b) Effect of inhibitors (10 mM) on enzyme activity (EDTA: Ethylenediaminetetraacetic acid, EGTA: Ethylene glycol tetraacetic acid, PCMB: p-chloromercuribenzoic acid, PMSF: Phenylmethanesulfonylfluoride, PBA c) Effect of serum and serum components (10 mM) on enzyme activity; d) Substrate (10 mM) specificity of enzyme.

<p>100% activity corresponds to 140 U of enzyme. Error bars represent SD of three experiments.</p

CD spectra of purified L-asparaginase a) Far UV CD spectra of L-asparaginase at 0.1 mg/ml in 0.1 M tris HCL (pH-8.4); b) Melting temperature of enzyme (T₂₂₂_nm).

<p>c). Near UV CD spectra of purified L-asparaginase 1.0 mg/ml in 0.1 M Tris HCL (pH-8.4).</p

Molecular weight determination of purified enzyme (a) SDS PAGE for steps of purifying L-asparaginase; b) Plot of V_e/V_o against semi-log of molecular weight of proteins on Sephacryl TM S-200 high resolution column (16/60) for α-Lactalbumin (12.4 kDa), carbonic anhydrase (30 kDa), bovine serum albumin (66 kDa), yeast alcohol dehydrogenase (150 kDa), sweet potato β-amylase (200 kDa) and ferritin (450 kDa).

<p>Molecular weight determination of purified enzyme (a) SDS PAGE for steps of purifying L-asparaginase; b) Plot of V_e/V_o against semi-log of molecular weight of proteins on Sephacryl TM S-200 high resolution column (16/60) for α-Lactalbumin (12.4 kDa), carbonic anhydrase (30 kDa), bovine serum albumin (66 kDa), yeast alcohol dehydrogenase (150 kDa), sweet potato β-amylase (200 kDa) and ferritin (450 kDa).</p

Effect of physical parameters on purified L-asparaginase a) Effect of pH on enzyme activity; b) Effect of pH on the stability of enzyme; c) Effect of temperature on assay reaction; d) Heat stability of enzyme.

<p>100% activity corresponds to 140 U of enzyme. Error bars represent SD of three experiments.</p