Purification of recombinant alpha-amylase by immunoaffinity chromatography with anti-peptide antibody.
Adsorption characteristics of an anti-peptide antibody, obtained by immunization of eight amino acids in the C-terminal region of chimeric alpha-amylase of rice alpha-amylase isozymes, were studied by use of the chimeric enzyme and the peptide used for immunization. This anti-peptide antibody adsorbed the enzyme, as well as the peptide antigen, with sufficient affinity for immunoaffinity purification and was used for purification of the enzyme secreted from yeast cells. Chimeric alpha-amylase was purified by immunoaffinity chromatography to high purity in one step from the fermentation broth. One-third of the secreted enzyme was not adsorbed by the column of anti-peptide antibody because of processing in the C-terminal region.
Katoh
S
S
Department of Synthetic Chemistry and Biological Chemistry, Kyoto University, Japan.
Terashima
M
M
Kouno
M
M
D000595:Amino Acid Sequence; D000906:Antibodies; D002846:Chromatography, Affinity / Q000379:methods*; D005285:Fermentation; D007527:Isoenzymes / Q000276:immunology; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000276:immunology; D012275:Oryza / Q000201:enzymology; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D000516:alpha-Amylases / Q000276:immunology / Q000302:isolation & purification*
D000906:Antibodies; D007527:Isoenzymes; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D000516:alpha-Amylases