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Purification of Recombinant Brain Derived Neurotrophic Factor Using Reversed Phase Displacement Chromatography
Author(s) -
Sunasara Khurram M.,
Rupp Randall G.,
Cramer Steven M.
Publication year - 2001
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp0100833
Subject(s) - trifluoroacetic acid , displacement chromatography , chromatography , yield (engineering) , chemistry , elution , impurity , displacement (psychology) , recombinant dna , ion chromatography , high performance liquid chromatography , reversed phase chromatography , materials science , biochemistry , organic chemistry , psychology , metallurgy , psychotherapist , gene
This work investigates the utility of RPLC displacement chromatography for the purification of recombinant brain derived neurotrophic factor (rHu‐BDNF) from its variants and E. coli. protein (ECP) impurities. The closely associated variants (six in total) differ by one amino acid from the native BDNF and thus pose a challenging separation problem. Several operational parameters were investigated to study their effects on the yield of the displacement process. The results indicated that the concentration of trifluoroacetic acid (TFA) in the buffer was a key factor in achieving the desired purification. Displacement chromatography on an analytical scale column resulted in extremely high purity and yield in a single chromatographic step. The process was successfully scaled‐up with respect to particle and column diameter. The production rate of a pilot scale RPLC displacement process was shown to be 23 times higher than the combined production rates of the current preparative ion exchange and hydrophobic interaction gradient elution steps that are used to remove variant and ECP impurities, respectively.