Improved purification process of β- and α-trypsin isoforms by ion-exchange chromatography

• Autor(a) principal: Santos,Alexandre Martins Costa
• Data de Publicação: 2008
• Outros Autores: Oliveira,Jamil Silvano de, Bittar,Eustáquio Resende, Silva,Anderson Lourenço da, Guia,Marcos Luiz dos Mares, Bemquerer,Marcelo Porto, Santoro,Marcelo Matos
• Tipo de documento: Artigo
• Idioma: eng
• Título da fonte: Brazilian Archives of Biology and Technology
• Texto Completo: http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132008000400009
• topic: Trypsin isoforms; resolution; ion-exchange chromatography; purification; specific activity; mass spectrometry

The purpose of this work was to improve the separation and yield of pure β- and α-trypsin isoforms by ion-exchange chromatography and to characterize some physical-chemical properties of these isoforms. Purification of trypsin isoforms was performed by ion-exchange chromatography in 0.1 mol/L tris-HC buffer, pH 7.10 at 4ºC. The sample loading, salt concentration, flow rate and pH of mobile phase were varied to determine their effects on the resolution of the separation. The resolution was optimized mainly between β- and α-trypsin. Pure isoforms were obtained by chromatographying 100 mg of commercial trypsin during seven days, yielding 51 mg of high purity β-trypsin and 13 mg of α-trypsin partially pure, with small amounts of contaminating of ψ-trypsin. Thus, time and resolution of purification were optimized yielding large amounts of pure active enzymes that are useful for several research areas and biotechnology.