Phastgel ief 3 9

Protein concentration was determined with a Quick Start Bradford protein assay kit (Bio-Rad Laboratories, Hercules, CA, USA). The apparent molecular mass of the enzymes was determined by SDS-PAGE in a 10–15 % polyacrylamide gel gradient as described by Laemmli. ¹⁷⁾ A BenchMark Ladder (Invitrogen, Carlsbad, CA, USA) provided the standard protein markers. The molecular mass of the native enzyme was determined by native-PAGE in an 8–25 % polyacrylamide gel gradient as described by Davis. ¹⁸⁾ The HMW marker kit (GE Healthcare, Chicago, IL, USA) provided the standard protein markers. The p I for thyroglobulin (669 kDa), ferritin (440 kDa), catalase (232 kDa), lactate dehydrogenase (140 kDa), and albumin (66 kDa) in the protein marker mix were 4.5, 4.5, 5.4, 5.0, and 4.9, respectively. The apparent p I of the enzyme was determined by isoelectric focusing on a Phastgel IEF 3–9 using the Phastsystem (GE Healthcare, Chicago, IL, USA). The proteins were stained with Rapid CBB KANTO (Kanto Chemical Co. Inc., Chuo-ku, Japan).

Protein concentration was determined using Quick Start Bradford Protein Assay (Bio-Rad, USA). The apparent molecular mass of the enzymes was determined using SDS-PAGE in 10–15 % gradient polyacrylamide gels as described by Laemmli.¹¹⁾ (link) Benchmark Ladder (Invitrogen) was used as standard protein markers. The molecular mass of the native enzymes was determined using native-PAGE in 8–25 % gradient polyacrylamide gels as described by Davis.¹²⁾ HMW Marker Kit (GE Healthcare) was used as standard protein markers. The pI values of thyroglobulin (669 kDa), ferritin (440 kDa), catalase (232 kDa), lactate dehydrogenase (140 kDa), and albumin (66 kDa) present in the protein marker are 4.5, 4.5, 5.4, 5.0, and 4.9, respectively. The apparent pI of the enzymes was determined using isoelectric focusing on a Phastgel IEF 3–9 and the Phastsystem (GE Healthcare). Proteins were stained using Rapid CBB KANTO (Kanto Chemical, Japan).