Fig. 4

SDS–PAGE and western blot analysis of recombinant His-tagged GH16B β-agarase with or without N-terminal 22-aa signal sequence. a, c Equivalent portions of individual intracellular fractions and culture supernatants of E. coli transformants, and GH16B β-agarase purified from the culture supernatants using ammonium sulfate precipitation and the Ni-NTA purification system were subjected to SDS-PAGE and the gels were stained with CBB R-250 to detect protein bands. b, d For western blot analysis, proteins in the gels were electrotransferred onto Immobilon-P membranes. The membrane was then probed with antibodies specific to the protein of interest. Protein detection was performed using the ECL western blotting detection system. Lane: SM, prestained size markers; GH16B-w/SS, GH16B β-agarase with the signal sequence; GH16B-w/oSS, GH16B β-agarase without the signal sequence; crude GH16B-w/SS, enzyme in the culture supernatants (CS); salting-out GH16B-w/SS, enzyme concentrated using ammonium sulfate salting-out of the CS; purified GH16B-w/SS, enzyme purified from the concentrated GH16B-w/SS using the Ni-NTA purification system