| <b>Unit Definition</b> One unit will digest urea-denatured hemoglobin at 37°C (pH 7.5) per minute to produce equal absorbance as 1.0 μmol of L-tyrosine using Folin & Ciocalteu's phenol reagent (6). <b>Storage Buffer</b> 20 mM Tris-HCl 1 mM CaCl2 50% Glycerol pH 7.4 @ 25°C <b>Heat Inactivation</b> 95°C for 10 minutes <b>Molecular Weight</b> Theoretical | 28.9 kDa <b>Unit Assay Conditions</b> 0.5–2 μg of Proteinase K is incubated with 2% denatured hemoglobin solution for 10 minutes at 37°C (pH 7.5). After precipitation, neutralization and addition of Folin & Ciocalteu's phenol reagent, absorbance of soluble cleavage products are measured at 750 nm. Absorbance is compared to a standard curve of L-tyrosine absorbance prepared similarly. |
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