Development of the purification process of pneumococcal surface protein A clade 4 (PspA4Pro).

Author(s)

Douglas Borges de Figueiredo

• Document type: Master's Dissertation
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Instituto de Ciências Biomédicas (ICB/SDI)
• Defense date: 2014-09-23

Examining board members

Viviane Maimoni Gonçalves; Adriano Rodrigues Azzoni; Giovana Cappio Barazzone

• Advisor: Viviane Maimoni Gonçalves; Joaquín Cabrera Crespo

Abstract

Pneumococcal surface protein A (PspA) is found in all Streptococcus pneumoniae strains and is a promising candidate to be used in new pneumococcal vaccines. A purification process for PspA4Pro which inicial steps were: cell disruption, precipitation of the homogenate with the cationic detergent cetyltrimethylammonium bromide (CTAB) and pellet removal by centrifugation. The chromatographic techniques tested were ion exchange (anionic and cationic), immobilized metal affinity, hydrophobic interaction and mix mode with hydrophobic and cationic ligands. Using CTAB precipitation, anion exchange chromatography, crioprecipitation in pH4.0 and cation exchange chromatography the PspA reached the required purity (>95%) with recovery between 14% and 33% . The process reached acceptable levels of endotoxin in the final product and the purified PspA4Pro was recognized by anti-PspA4 antibodies and manteined its activity and secondary structure. (AU)

• FAPESP's process: 12/04858-7 - Development of the purification process of the pneumococcal surface protein a from Clade4 (PspA4pro)
• Grantee: Douglas Borges de Figueiredo
• Support Opportunities: Scholarships in Brazil - Master